U.S. patent application number 15/865004 was filed with the patent office on 2018-08-09 for devices, systems and methods for sample preparation.
The applicant listed for this patent is Theranos, Inc.. Invention is credited to Elizabeth A. Holmes, Alexander Loo.
Application Number | 20180224472 15/865004 |
Document ID | / |
Family ID | 51580735 |
Filed Date | 2018-08-09 |
United States Patent
Application |
20180224472 |
Kind Code |
A1 |
Loo; Alexander ; et
al. |
August 9, 2018 |
DEVICES, SYSTEMS AND METHODS FOR SAMPLE PREPARATION
Abstract
Devices, systems and methods including a sonicator for sample
preparation are provided. A sonicator may be used to mix,
resuspend, aerosolize, disperse, disintegrate, or de-gas a
solution. A sonicator may be used to disrupt a cell, such as a
pathogen cell in a sample. Sample preparation may include exposing
pathogen-identifying material by sonication to detect, identify, or
measure pathogens. A sonicator may transfer ultrasonic energy to
the sample solution by contacting its tip to an exterior wall of a
vessel containing the sample. Multipurpose devices including a
sonicator also include further components for additional actions
and assays. Devices, and systems comprising such devices, may
communicate with a laboratory or other devices in a system for
sample assay and analysis. Methods utilizing such devices and
systems are provided. The improved sample preparation devices,
systems and methods are useful for analyzing samples, e.g. for
diagnosing patients suffering from infection by pathogens.
Inventors: |
Loo; Alexander; (Redwood
City, CA) ; Holmes; Elizabeth A.; (Palo Alto,
CA) |
|
Applicant: |
Name |
City |
State |
Country |
Type |
Theranos, Inc. |
Newark |
CA |
US |
|
|
Family ID: |
51580735 |
Appl. No.: |
15/865004 |
Filed: |
January 8, 2018 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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14845740 |
Sep 4, 2015 |
9885729 |
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15865004 |
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PCT/US14/22847 |
Mar 10, 2014 |
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14845740 |
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61806809 |
Mar 29, 2013 |
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61799533 |
Mar 15, 2013 |
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Current U.S.
Class: |
1/1 |
Current CPC
Class: |
G01N 1/38 20130101; G01N
1/28 20130101; B01F 11/0266 20130101; G01N 35/1009 20130101; G01N
35/00 20130101; G01N 33/487 20130101; G01N 35/00871 20130101; C12Q
1/04 20130101; G01N 1/286 20130101; G01N 2001/4094 20130101; G01N
2035/00554 20130101; G01N 2035/00524 20130101 |
International
Class: |
G01N 35/00 20060101
G01N035/00; G01N 33/487 20060101 G01N033/487; C12Q 1/04 20060101
C12Q001/04; G01N 1/28 20060101 G01N001/28; G01N 1/38 20060101
G01N001/38; B01F 11/02 20060101 B01F011/02 |
Claims
1.-39. (canceled)
40. A method of detecting the presence of pathogen-identifying
material in a biological sample, comprising: transporting at least
a portion of a biological sample to a vessel, said vessel having a
mating socket, an interior portion, and an exterior wall, effective
to place said portion of said biological sample into said interior
portion of said vessel; contacting said mating socket with a sample
handling system effective to engage said vessel with said sample
handling system; transporting said vessel by said sample handling
system to a location adjacent a sonicator; contacting an exterior
wall of said vessel containing said biological sample with said
sonicator; applying ultrasound energy to said wall of the vessel
effective to release pathogen-identifying material from a target
pathogen in said biological sample; and detecting the presence of
pathogen-identifying material.
41. The method of detecting the presence of pathogen-identifying
material of claim 40, wherein said detecting the presence of
pathogen-identifying material comprises performing an isothermal
assay.
42. The method of detecting the presence of pathogen-identifying
material in a biological sample of claim 40, wherein said
biological sample comprises a sample selected from blood, urine,
sputum, tears, material from a nasal swab, material from a throat
swab, material from a cheek swab, and another bodily fluid,
excretion, secretion, and tissue obtained from a subject.
43. A method of measuring an amount of a pathogen-identifying
material in a biological sample, comprising: Contacting said
biological sample with an automated assay device; and Measuring an
amount of a pathogen-identifying material in said sample. wherein
said automated assay device comprises a sample handling system; a
detector, a vessel holder configured to hold a vessel having a
vessel wall, said vessel wall having an external face, said vessel
being configured to hold a sample; and a sonicator, wherein said
sonicator is configured to contact said external face of said
vessel wall effective to disrupt a cell in a sample within said
vessel.
44. The method of measuring an amount of a pathogen-identifying
material of claim 43, wherein said measuring an amount of
pathogen-identifying material comprises performing an isothermal
assay.
45. The method of measuring an amount of a pathogen-identifying
material of claim 44, wherein said contacting step comprises:
transporting at least a portion of said biological sample to a
vessel, said vessel having an interior portion and a vessel wall
having an external face, effective to place said portion of said
biological sample into said interior portion of said vessel;
contacting said external face of said vessel wall with a sonicator;
and applying ultrasound energy to said vessel wall effective to
release pathogen-identifying material from a target pathogen in
said biological sample.
46. The method of claim 44, wherein said measuring comprises
optical detection.
47. The method of measuring an amount of a pathogen-identifying
material in a biological sample of claim 44, wherein said
biological sample comprises a sample selected from blood, urine,
sputum, tears, material from a nasal swab, material from a throat
swab, material from a cheek swab, and another bodily fluid,
excretion, secretion, and tissue obtained from a subject.
48. A method of measuring an amount of a pathogen-identifying
material in a biological sample, comprising: transporting at least
a portion of a biological sample to a vessel, said vessel having a
mating socket, an interior portion, and an exterior wall, effective
to place said portion of said biological sample into said interior
portion of said vessel; contacting said mating socket with a sample
handling system effective to engage said vessel with said sample
handling system; transporting said vessel by said sample handling
system to a location adjacent a sonicator; contacting an exterior
wall of said vessel containing said biological sample with said
sonicator; applying ultrasound energy to said wall of the vessel
effective to release pathogen-identifying material from a target
pathogen in said biological sample; and measuring the amount of a
pathogen-identifying material.
49-67. (canceled)
Description
CROSS-REFERENCE TO RELATED APPLICATIONS
[0001] This application claims priority to, and the benefit under
35 U.S.C. .sctn..sctn. 120 and 365(c), of international application
PCT/US2014/022847, filed Mar. 10, 2014, which international
application claims priority to, and the benefit under 35 U.S.C.
.sctn. 119(e) of, U.S. Patent Application 61/799,533, filed Mar.
15, 2013, and U.S. Patent Application 61/806,809, filed Mar. 29,
2013, all of which applications are hereby incorporated by
reference in their entireties.
BACKGROUND
[0002] Rapid and accurate detection and identification of pathogens
such as bacteria is important, and can be critical, in the
diagnosis and treatment of infectious diseases, and in the
prevention or amelioration of pandemics or epidemics. However,
methods for detecting or identifying pathogens often require large
samples, which can be difficult to obtain, or may be uncomfortable
or painful for the subject. Methods for detecting or identifying
pathogens often require the incubation of samples on growth media,
or in host cells or animals, and so take long periods of time. Many
pathogens are difficult to culture, or are similar to other
organisms, and so are difficult to identify even if the sample
yields a detectable culture. In addition, methods for detecting or
identifying pathogens may require rare or expensive reagents or
culture conditions.
[0003] Thus, detection and identification of pathogens in a
biological sample may be important in the diagnosis and treatment
of patients exposed to, or suspected of suffering, from infectious
diseases. However, present methods for detecting or identifying
pathogens are often inaccurate, difficult, expensive, and
time-consuming. Accordingly, rapid, accurate and straightforward
methods for the detection and identification of pathogens from
small samples are desired.
INCORPORATION BY REFERENCE
[0004] All publications, patents, and patent applications mentioned
in this specification are herein incorporated by reference to the
same extent as if each individual publication, patent, or patent
application was specifically and individually indicated to be
incorporated by reference.
SUMMARY
[0005] Devices, systems and methods for sample preparation are
disclosed. Devices for sample preparation as disclosed herein
comprise a sonicator; the sonicator may be configured to provide
ultrasonic energy to a sample solution. A sample solution may be
contained within a vessel. In embodiments, a sonicator may be used
to mix a solution, e.g., to mix a sample solution, or to mix two or
more solutions, such as a sample solution and a reagent solution.
In embodiments, a sonicator may be used for emulsification of a
solution or a mixture of solutions, or of a sample in a solution or
reagent. In embodiments, a sonicator may be used for resuspension
of a material, e.g., resuspension of a sample following
centrifugation of the sample. In embodiments, a sonicator may be
used for aerosolization of a liquid, e.g., aerosolization of a
liquid sample, or of a solution comprising a sample. In
embodiments, a sonicator may be used to heat a solution, such as a
sample, or a solution comprising a sample. In embodiments, a
sonicator may be used for dispersing a sample, or a sample in a
solution, or other material, such as, e.g., dispersing a solid or
semi-solid sample in a solution. In embodiments, a sonicator may be
used to disintegrate a material, such as a sample, e.g., a solid or
semi-solid sample may be exposed to sonication for disintegration,
which may aid in its subsequent mixing into a reagent such as a
diluent. In embodiments, a sonicator may be used to de-gas a
liquid, such as a fluid sample; in embodiments, gas released in
this way may become better available for assay. In embodiments, a
sonicator may be used to disrupt a cell, such as a pathogen cell in
a sample. Disruption of a cell will typically expose cellular
contents (intracellular material, including, e.g., nuclear material
from within a cell or organelle) to testing, and to provide more
accessible proteins and protein/membrane complexes for assay
[0006] Pathogens may be detected and identified by their
characteristic genetic material and characteristic proteins.
Detection and identification of pathogens in a biological sample
typically requires disruption of the pathogen to expose its nuclear
material to testing, and to provide more accessible proteins and
protein/membrane complexes for assay. Pathogen-identifying material
may be released from the pathogen by disruption, e.g., by
disruption of a pathogen membrane or cell wall. For example,
disruption of the plasma membrane of any cell, or of a nuclear
membrane of a eukaryotic pathogen, or disruption of the cell wall
of a bacterium having a cell wall, or a combination of the same,
may be effective to free pathogen-identifying material (e.g., DNA,
RNA, or protein) for contact with assay reagents for detection,
identification, or measurement.
[0007] Devices, systems and methods for disrupting pathogens in a
biological sample are provided. Devices, systems and methods for
disrupting pathogens in a biological sample as disclosed herein
comprise a sonicator; the sonicator may be configured to provide
ultrasonic energy to a sample solution containing a pathogen
effective to disrupt the pathogen, e.g., to disrupt a membrane of
the pathogen. Such disruption may be effective to release
pathogen-identifying material from the pathogen, aiding in the
detection, identification, or measurement of the pathogen. In
embodiments, multipurpose devices are provided which include a
sonicator configured to disrupt pathogen cells in a biological
sample, exposing pathogen-identifying material for assay.
Multipurpose devices are provided which include a sonicator
configured to disrupt pathogen cells in a biological sample,
exposing pathogen-identifying material for assay, and which may
also perform at least one additional assay are provided. Systems
comprising such devices are provided. Systems comprising such
devices may further communicate with a laboratory or with other
devices in a system for the detection and identification of
pathogens, and for diagnosis of infectious diseases. Methods
utilizing such devices and systems are provided. The devices,
systems and methods disclosed herein provide advantages useful for
diagnosing and treating patients suspected of, or suffering from,
infection by pathogens. A biological sample may comprise blood,
urine, sputum, tears, material obtained from a nasal swab, throat
swab, cheek swab, or other sample obtained from a subject.
[0008] Applicants disclose devices configured to assay a biological
sample for the presence of pathogen-identifying material, said
devices comprising a sonicator, said sonicator being configured to
contact a vessel containing said biological sample. Such a
sonicator may be configured for use in the performance of said
assay. A sonicator in a device as disclosed herein may be
configured to contact a wall of a vessel containing at least a
portion of a biological sample, effective to provide ultrasonic
energy to said wall, effective to disrupt a pathogen in said
biological sample and to release pathogen-identifying material from
said pathogen. An assay may comprise detection of
pathogen-identifying material; an assay may comprise identification
of pathogen-identifying material; an assay may comprise measuring
pathogen-identifying material, e.g., measuring an amount of
pathogen-identifying material in a sample or portion of a sample.
Such pathogen-identifying material may be detected, or may be
identified, or may be measured, in an assay performed with said
biological sample. In embodiments, a device disclosed herein may be
configured to perform one or more other assay in addition to an
assay directed to the detection, identification, or measurement of
pathogen-identifying material.
[0009] A device as disclosed herein may be suitable for detection,
identification, or measurement of pathogen-identifying material in
a biological sample. In embodiments, a device for assaying a sample
for the presence of pathogen-identifying material in a sample may
comprise a sonicator, wherein said sonicator comprises means for
contacting a wall of a vessel containing at least a portion of a
biological sample. In embodiments, a device for assaying a sample
for the presence of pathogen-identifying material in a sample may
comprise: a sonicator; an sample handling system for transporting
at least a portion of a biological sample; a vessel holder
effective to hold a vessel having a wall; and a detector effective
to detect or measure pathogen-identifying material (e.g., an
optical detector). In embodiments, a sonicator may comprise a tip;
in embodiments, a sonicator tip may be effective to contact said
vessel wall and to transfer ultrasonic energy to said vessel wall
from said sonicator. In embodiments, a sonicator may comprise a
sonicator horn, which horn may have a tip, and said horn and tip
may be effective to contact said vessel wall and to transfer
ultrasonic energy to said vessel wall from said sonicator. In
embodiments, a device may further comprise a communication
assembly, which may comprise a display element or a communication
element effective to report the results of said detection and/or
measurement. In embodiments, a communication assembly, such as a
display element and/or communication element, may be suitable for
two-way communication.
[0010] Applicants disclose herein an automated assay device
configured to disrupt a cell in a sample, the device comprising a
sample handling system; a detector, a vessel holder configured to
hold a vessel having a vessel wall, the vessel wall having an
external face, the vessel being configured to hold a sample; and a
sonicator, wherein the sonicator is configured to contact the
external face of the vessel wall effective to disrupt a cell in a
sample within the vessel. In embodiments, such an automated assay
device is configured to perform an assay on a sample when the
sample is held in a vessel held by a vessel holder, the assay
comprising an assay for pathogen-identifying material in the
sample. In embodiments, a sonicator of such an automated assay
device has a sonicator tip configured to contact the external face
of the vessel wall effective to transfer ultrasonic energy from the
sonicator to the vessel wall upon operation of the sonicator. In
embodiments, a detector of such an automated assay device includes
an optical detector. In embodiments, the device is configured to
apply force to a vessel while applying a sonicator tip to a wall of
a vessel.
[0011] An automated assay device as disclosed herein may further
include a communication assembly configured to communicate with a
user, a device, a laboratory, a network, or the cloud.
[0012] The assay for pathogen-identifying material of such an
automated assay device may be or include an assay selected from an
assay for the detection of pathogen-identifying material in a
sample; an assay for the identification of pathogen-identifying
material in a sample; and an assay for the measurement of the
amount of pathogen-identifying material in a sample. In
embodiments, the assay for pathogen-identifying material comprises
an isothermal assay.
[0013] In embodiments, an automated assay device configured to
assay a sample for pathogen-identifying material comprises a sample
handling means; a vessel holding means configured to hold a vessel
having a vessel wall having an external face; a detector means; and
a sonicator; wherein the sonicator is configured to contact the
external face of the vessel wall effective to transfer ultrasonic
energy from the sonicator to the vessel wall upon operation of the
sonicator. In embodiments, the detector means comprises optical
detection means. In embodiments, an assay of such an automated
assay device comprises an assay selected from an assay for
detection pathogen-identifying material in a sample; an assay for
identification of pathogen-identifying material in a sample; and an
assay for measurement of an amount of pathogen-identifying material
in a sample. In embodiments, the assay for pathogen-identifying
material comprises an isothermal assay.
[0014] Biological samples suitable for use in the devices disclosed
herein, and suitable for assay by an automated assay device as
disclosed herein, include samples selected from blood, urine,
sputum, tears, material from a nasal swab, material from a throat
swab, material from a cheek swab, and another bodily fluid,
excretion, secretion, and tissue obtained from a subject.
[0015] Applicants also disclose herein methods of detecting the
presence of pathogen-identifying material in a biological sample.
Biological samples suitable for use in such methods include samples
selected from blood, urine, sputum, tears, material from a nasal
swab, material from a throat swab, material from a cheek swab, and
another bodily fluid, excretion, secretion, and tissue obtained
from a subject.
[0016] For example, Applicants disclose a method of detecting the
presence of pathogen-identifying material in a biological sample,
comprising: [0017] Contacting the biological sample with an
automated assay device as disclosed herein; and [0018] Detecting
the presence of a pathogen-identifying material in the sample.
[0019] In embodiments of such a method, the contacting step may
comprise: [0020] transporting at least a portion of the biological
sample to a vessel, the vessel having an interior portion and a
vessel wall having an external face, effective to place the portion
of the biological sample into the interior portion of the vessel;
[0021] contacting the external face of the vessel wall with a
sonicator; and [0022] applying ultrasound energy to the vessel wall
effective to release pathogen-identifying material from a target
pathogen in the biological sample.
[0023] The detecting in such methods may comprise optical
detection.
[0024] In embodiments, a method of detecting the presence of
pathogen-identifying material as disclosed herein comprises
performing an isothermal assay for the detection of the presence of
pathogen-identifying material.
[0025] A further method of identifying pathogen-identifying
material in a biological sample, as disclosed herein, comprises:
[0026] Contacting the biological sample with an automated assay
device having features as disclosed herein; and [0027] Identifying
pathogen-identifying material in the sample.
[0028] In embodiments, the contacting step comprises: [0029]
transporting at least a portion of the biological sample to a
vessel, the vessel having an interior portion and a vessel wall
having an external face, effective to place the portion of the
biological sample into the interior portion of the vessel; [0030]
contacting the external face of the vessel wall with a sonicator;
and applying ultrasound energy to the vessel wall effective to
release pathogen-identifying material from a target pathogen in the
biological sample.
[0031] In embodiments, identifying comprises optical detection.
[0032] In embodiments of the methods disclosed herein, the
identification of pathogen-identifying material comprises
performing an isothermal assay for the identification of
pathogen-identifying material.
[0033] Applicants further disclose a method of disrupting a
pathogen in a biological sample, comprising: [0034] transporting at
least a portion of a biological sample to a vessel, the vessel
having an interior portion and a vessel wall having an external
face, effective to place the portion of the biological sample into
the interior portion of the vessel; [0035] contacting the external
face of the vessel with a sonicator; and [0036] applying ultrasound
energy to the wall of the vessel effective to disrupt a target
pathogen in the biological sample.
[0037] A yet further method of detecting the presence of
pathogen-identifying material in a biological sample comprises:
[0038] transporting at least a portion of a biological sample to a
vessel, the vessel having a mating socket, an interior portion, and
an exterior wall, effective to place the portion of the biological
sample into the interior portion of the vessel; [0039] contacting
the mating socket with a sample handling system effective to engage
the vessel with the sample handling system; [0040] transporting the
vessel by the sample handling system to a location adjacent a
sonicator; [0041] contacting an exterior wall of the vessel
containing the biological sample with the sonicator; [0042]
applying ultrasound energy to the wall of the vessel effective to
release pathogen-identifying material from a target pathogen in the
biological sample; and detecting the presence of
pathogen-identifying material.
[0043] In embodiments, the detecting the presence of
pathogen-identifying material in such a method comprises performing
an isothermal assay. In embodiments, the detecting the presence of
pathogen-identifying material in such a method comprises optical
detection.
[0044] Another method of measuring an amount of a
pathogen-identifying material in a biological sample, comprising:
[0045] Contacting the biological sample with an automated assay
device disclosed herein; and [0046] Measuring an amount of a
pathogen-identifying material in the sample.
[0047] In embodiments, measuring an amount of pathogen-identifying
material comprises performing an isothermal assay.
[0048] In embodiments, the contacting step of such a method
comprises: [0049] transporting at least a portion of the biological
sample to a vessel, the vessel having an interior portion and a
vessel wall having an external face, effective to place the portion
of the biological sample into the interior portion of the vessel;
[0050] contacting the external face of the vessel wall with a
sonicator; and [0051] applying ultrasound energy to the vessel wall
effective to release pathogen-identifying material from a target
pathogen in the biological sample.
[0052] In embodiments, the measuring step comprises optical
detection.
[0053] Another method of measuring an amount of a
pathogen-identifying material in a biological sample comprises:
[0054] transporting at least a portion of a biological sample to a
vessel, the vessel having a mating socket, an interior portion, and
an exterior wall, effective to place the portion of the biological
sample into the interior portion of the vessel; [0055] contacting
the mating socket with a sample handling system effective to engage
the vessel with the sample handling system; [0056] transporting the
vessel by the sample handling system to a location adjacent a
sonicator; [0057] contacting an exterior wall of the vessel
containing the biological sample with the sonicator; [0058]
applying ultrasound energy to the wall of the vessel effective to
release pathogen-identifying material from a target pathogen in the
biological sample; and [0059] measuring the amount of a
pathogen-identifying material.
[0060] Applicants also disclose systems comprising an automated
assay device as disclosed herein. In embodiments, such a system
comprises an automated assay device as disclosed herein, wherein
the automated assay device comprises a sonicator having a sonicator
tip; and a vessel, wherein the vessel comprises a vessel wall
having an external face, the external face comprising a
sonicator-contacting portion, wherein the sonicator-contacting
portion comprises a flat area complementary to a flat portion of
the sonicator tip.
[0061] The flat area of the vessel comprises a flat area of a side
wall of the vessel in embodiments of such a system. In embodiments,
the flat area of a vessel comprises a flat area of a bottom wall of
the vessel. In embodiments, the vessel comprises a plurality of
flat areas; in embodiments, at least one of the flat areas of the
vessel comprises a flat area of a side wall of the vessel; or at
least one of the flat areas of the vessel comprises a flat area of
a bottom wall of the vessel; orat least one of the flat areas of
the vessel comprises a flat area of a side wall of the vessel, and
at least one of the flat areas of the vessel comprises a flat area
of a bottom wall of the vessel.
[0062] Applicants further disclose vessels herein, such vessels
comprising an internal portion for holding a sample, a side wall, a
bottom wall, and a mating socket configured to mate with a sample
handling system, wherein at least one of the vessel walls comprises
a flat area configured to make contact with a sonicator tip
effective that ultrasonic energy from a sonicator in contact with
the vessel is transferred to a sample held in the internal portion
during operation of the sonicator. In embodiments, the flat area of
the vessel comprises a flat area of a side wall of the vessel; or
the flat area of the vessel comprises a flat area of a bottom wall
of the vessel; or the vessel comprises a plurality of flat areas;
or at least one of the flat areas of the vessel comprises a flat
area of a side wall of the vessel; or at least one of the flat
areas of the vessel comprises a flat area of a bottom wall of the
vessel; or at least one of the flat areas of the vessel comprises a
flat area of a side wall of the vessel, and at least one of the
flat areas of the vessel comprises a flat area of a bottom wall of
the vessel; and combinations and multiples thereof.
[0063] In embodiments, a device for assaying a sample for the
presence of pathogen-identifying material in a sample may comprise
a sonicator, wherein said sonicator comprises means for contacting
a wall of a vessel containing at least a portion of a biological
sample. In embodiments, a device for assaying a sample for the
presence of pathogen-identifying material in a sample may comprise:
a sonicator; sample handling system means for transporting at least
a portion of a biological sample; means for holding a vessel having
a wall; means for contacting said vessel wall with at least a
portion of said sonicator; and means for detecting, identifying, or
measuring pathogen-identifying material (e.g., means for detecting
an optical signal, and/or measuring an optical property of the
combined composition with sample). In embodiments, a device may
further comprise means for displaying or reporting the results of
said detection and/or measurement. In embodiments, a sonicator may
comprise a tip means, which may comprise a sonicator horn,
configured to make effective contact with a surface, such as a
vessel wall, effective to transfer ultrasonic energy from said
sonicator to said surface. In further embodiments, a device for
assaying a sample for the presence of pathogen-identifying material
in a sample may comprise one or more of: detection means (e.g., an
optical detector) for detecting the target; display means; and
communication means for reporting the results of said detection
and/or measurement. In embodiments, display means and/or
communication means may be suitable for two-way communication.
[0064] Applicants further disclose systems comprising a device as
disclosed herein. In embodiments, a system as disclosed herein may
be used to detect the presence of, identify, or measure the amount
of, pathogen-identifying material in a sample.
[0065] Applicants disclose herein systems for detecting the
presence of pathogen-identifying material in a biological sample,
e.g., a sample of blood, urine, sputum, tears, material from a
nasal swab, throat swab, cheek swab, or other sample obtained from
a subject. In embodiments, a system for detecting the presence of
pathogen-identifying material in a sample may comprise a device as
disclosed herein, and a means for communicating information from
said device to a computer, a computer network, a telephone, a
telephone network, or a device configured to display information
communicated from said device.
[0066] Applicants disclose herein systems for identifying
pathogen-identifying material in a biological sample, e.g., a
sample of blood, urine, sputum, tears, material from a nasal swab,
throat swab, cheek swab, or other sample obtained from a subject.
In embodiments, a system for identifying pathogen-identifying
material in a sample may comprise a device as disclosed herein, and
a means for communicating information from said device to a
computer, a computer network, a wide-area network, a local-area
network, a virtual private network, a fiber-optical network, a
telephone, a telephone network (which may include both land-line
telephone networks and mobile telephone networks), or a device
configured to display information communicated from said
device.
[0067] Applicants disclose herein systems for measuring the amount
of pathogen-identifying material in a biological sample, e.g. a
sample of blood, urine, sputum, tears, material from a nasal swab,
throat swab, cheek swab, or other sample obtained from a subject.
In embodiments, a system for measuring the amount of
pathogen-identifying material in a sample may comprise a device as
disclosed herein, and a means for communicating information from
said device to a computer, a computer network, a telephone, a
telephone network, or a device configured to display information
communicated from said device.
[0068] It will be understood that a means for communicating
information may include means for one-way communication and may
include means for two-way communication, and may include means for
communication with multiple locations or entities. In embodiments,
a system for detecting the presence of pathogen-identifying
material in a sample, a system for identifying pathogen-identifying
material in a sample, and a system for measuring the amount of
pathogen-identifying material in a sample, may comprise a device as
disclosed herein, and a communication assembly, which may comprise
a channel for communicating information from said device to a
computer, said wherein said channel is selected from a computer
network, a telephone network, a metal communication link, an
optical communication link, and a wireless communication link. It
will be understood that a communication assembly, e.g., comprising
a channel for communicating information, may comprise a one-way
communication channel and may be a two-way communication channel,
and may comprise channels for communication with multiple locations
or entities.
[0069] The methods disclosed herein may be performed on a device,
or on a system, for processing a sample as disclosed herein. The
methods disclosed herein can be readily incorporated into and used
in an automated assay device, and in an automated assay system, as
disclosed herein. For example, systems as disclosed herein may
include a communication assembly for transmitting or receiving a
protocol based on the analyte to be detected (e.g.,
pathogen-identifying material) or based on other analytes to be
detected by the device or system. In embodiments, an assay protocol
may be changed based on optimal scheduling of a plurality of assays
to be performed by a device, or may be changed based on results
previously obtained from a sample from a subject, or based on
results previously obtained from a different sample from the
subject. In embodiments, a communication assembly may comprise a
channel for communicating information from said device to a
computer, said wherein said channel is selected from a computer
network, a telephone network, a metal communication link, an
optical communication link, and a wireless communication link. In
embodiments, systems as disclosed herein may transmit signals to a
central location, or to an end user, and may include a
communication assembly for transmitting such signals. Systems as
disclosed herein may be configured for updating a protocol as
needed or on a regular basis.
[0070] Accordingly, Applicants disclose devices configured to
detect, identify, or measure pathogen-identifying material in a
biological sample, e.g., of blood, urine, sputum, material obtained
from a nasal swab, a throat swab, a cheek swab, or other sample.
Such detection, identification, or measurements may be made
according to a method disclosed herein. Devices configured to
detect, identify, or measure pathogen-identifying material in a
biological sample, e.g., of blood, urine, sputum, material obtained
from a nasal swab, a throat swab, a cheek swab, or other sample,
according to a method disclosed herein may be configured to
determine pathogen-identifying material from a biological sample
that comprises no more than about 1000 .mu.L of a biological
sample, or no more than about 500 .mu.L of a biological sample, or
no more than about 250 .mu.L of a biological sample, or no more
than about 150 .mu.L of a biological sample, or no more than about
100 .mu.L of a biological sample, or no more than about 50 .mu.L of
a biological sample, or, in embodiments, wherein said sample of
blood comprises no more than about 25 .mu.L of a biological sample,
or wherein said sample of blood comprises no more than about 10
.mu.L of a biological sample, or wherein said sample of blood
comprises less than about 10 .mu.L of a biological sample. Such
devices may be configured to detect, identify, or measure
pathogen-identifying material in a biological sample, e.g., of
blood, urine, sputum, material obtained from a nasal swab, a throat
swab, a cheek swab, or other sample, in less than about one hour,
or, in embodiments, in less than about 40 minutes, or in less than
about 30 minutes, or in less than about 20 minutes, or less than
about 10 minutes, or less than about 5 minutes, or less.
[0071] Devices disclosed herein may be configured to perform an
assay for the detection, identification, or measurement of
pathogen-identifying material, and also to perform an assay for the
measurement of another analyte in the biological sample, e.g., of
blood, urine, sputum, material obtained from a nasal swab, a throat
swab, a cheek swab, or other sample. Devices disclosed herein may
be configured to perform an assay for the detection,
identification, or measurement of pathogen-identifying material,
and also to perform an assay comprising measurement of a
morphological characteristic of a blood cell in the biological
sample, e.g., of blood, urine, sputum, material obtained from a
nasal swab, a throat swab, a cheek swab, or other sample. Devices
disclosed herein may be configured to perform an assay for the
detection, identification, or measurement of pathogen-identifying
material, and also to perform an assay comprising measurement of
another analyte, e.g., a vitamin, a hormone, a drug or metabolite
of a drug, or other analyte. Such devices may be configured wherein
the assays, or the order of performance of assays, that are
performed by said device may be altered by communication with
another device.
[0072] Applicants also disclose systems comprising a device as
disclosed herein. In embodiments, the system comprises a device
that is configured to perform an assay for the measurement of
pathogen-identifying material and also to perform an assay for the
measurement of another analyte in the biological sample, e.g., of
blood, urine, sputum, material obtained from a nasal swab, a throat
swab, a cheek swab, or other sample. In embodiments, the system
comprises a device that is configured to perform an assay for the
measurement of pathogen-identifying material and also to perform an
assay for the measurement of a morphological characteristic of a
blood cell in the biological sample, e.g., of blood, urine, sputum,
material obtained from a nasal swab, a throat swab, a cheek swab,
or other sample. In embodiments of such a system, assays, or the
order of performance of assays, that are performed by said device
may be altered by communication with another device.
[0073] Applicants disclose herein a method of disrupting a pathogen
in a biological sample, comprising: transporting at least a portion
of a biological sample to a vessel, said vessel having an interior
portion and an exterior wall, effective to place said portion of
said biological sample into said interior portion of said vessel;
contacting an exterior wall of said vessel containing said
biological sample with a sonicator; and applying ultrasound energy
to said wall of the vessel effective to disrupt a target pathogen
in said biological sample. In embodiments, such transporting of at
least a portion of a biological sample to a vessel may comprise
transporting by sample handling system.
[0074] Applicants disclose herein a method of detecting the
presence of pathogen-identifying material in a biological sample,
comprising: transporting at least a portion of a biological sample
to a vessel, said vessel having an interior portion and an exterior
wall, effective to place said portion of said biological sample
into said interior portion of said vessel; contacting an exterior
wall of said vessel containing said biological sample with a
sonicator; applying ultrasound energy to said wall of the vessel
effective to release pathogen-identifying material from a target
pathogen in said biological sample; and detecting the presence of
pathogen-identifying material. In embodiments, such transporting of
at least a portion of a biological sample to a vessel may comprise
transporting at least a portion of a biological sample by a sample
handling system.
[0075] Applicants disclose herein a method of identifying a
pathogen in a biological sample, comprising: transporting at least
a portion of a biological sample to a vessel, said vessel having an
interior portion and an exterior wall, effective to place said
portion of said biological sample into said interior portion of
said vessel; contacting an exterior wall of said vessel containing
said biological sample with a sonicator; applying ultrasound energy
to said wall of the vessel effective to disrupt a target pathogen
in said biological sample; and identifying a pathogen in the
sample. In embodiments, such transporting of at least a portion of
a biological sample to a vessel may comprise transporting at least
a portion of a biological sample by a sample handling system.
[0076] Applicants disclose herein a method of measuring the amount
of pathogen-identifying material in a biological sample,
comprising: transporting at least a portion of a biological sample
to a vessel, said vessel having an interior portion and an exterior
wall, effective to place said portion of said biological sample
into said interior portion of said vessel; contacting an exterior
wall of said vessel containing said biological sample with a
sonicator; applying ultrasound energy to said wall of the vessel
effective to release pathogen-identifying material from a target
pathogen in said biological sample; and measuring the amount of
pathogen-identifying material. In embodiments, such transporting of
at least a portion of a biological sample to a vessel may comprise
transporting at least a portion of a biological sample by a sample
handling system.
[0077] Applicants disclose herein devices and systems comprising a
sonicator, and methods using such device and systems to detect,
identify, or measure pathogens in a sample by detecting,
identifying, or measuring pathogen-identifying material released in
a sample as a result of disruption caused by application of
ultrasonic energy to the sample. As disclosed herein, ultrasonic
energy may be applied to a sample by direct contact between the
sample and a sonicator (e.g., between a sample and a tip portion of
a sonicator); by indirect contact between a sample and a sonicator,
wherein the sonicator contacts a compliant barrier that is in
contact with a sample; by indirect contact between a sample and a
sonicator, wherein the sonicator contacts a wall of a vessel
containing a sample; or by other means. Ultrasonic energy suitable
for disrupting pathogens using devices, systems, and methods as
disclosed herein includes ultrasonic energy delivered at between
about 20 kiloHertz (kHz) and about 60 kHz; between about 20 kHz and
about 50 kHz; between about 30 kHz and about 50 kHz; between about
20 kHz and about 40 kHz; or between about 20 kHz and about 30 kHz.
Ultrasonic energy suitable for disrupting pathogens using devise,
systems, and methods as disclosed herein includes ultrasonic energy
delivered at or about 20 kHz; at or about 25 kHz; at or about 28
kHz; at or about 30 kHz; at or about 35 kHz; at or about 40 kHz; at
or about 45 kHz; at or about 50 kHz; at or about 55 kHz; or at or
about 60 kHz.
[0078] Applicants disclose herein vessels; these vessels comprise
an internal portion for holding a sample, a side wall, and a bottom
wall. Some vessels disclosed herein comprise an internal portion
for holding a sample, a side wall, a bottom wall, and a mating
socket configured to mate with a sample handling system. Vessels
disclosed herein comprise at least one vessel wall which comprises
a flat area configured to make contact with a sonicator tip
effective that ultrasonic energy from a sonicator in contact with
the vessel is transferred to a sample held in said internal portion
during operation of a sonicator.
[0079] In embodiments, a flat area of a vessel comprises a flat
area of a side wall of the vessel. In embodiments, a flat area of a
vessel comprises a flat area of a bottom wall of the vessel. In
embodiments, a vessel comprises a plurality of flat areas. In
embodiments, at least one of the flat areas of a vessel comprises
flat area of a side wall of the vessel. In embodiments, at least
one of the flat areas of a vessel comprises a flat area of a bottom
wall of the vessel. In embodiments of vessels having a plurality of
flat areas, at least one flat area of a vessel comprises a flat
area of a side wall of the vessel, and at least one flat area
comprises a flat area of a bottom wall of the vessel.
[0080] In embodiments, a vessel is configured for the performance
of an optical assay for detection, identification, measurement, or
other characterization of a sample held within the vessel. In
embodiments, a vessel is transparent. In embodiments, a vessel is
translucent. A vessel may comprise a glass, a plastic, a ceramic, a
polymer, or other material, or combination of materials. A vessel
may comprise two or more materials; for example, a flat area may
comprise a different material than another portion of a vessel. A
vessel may comprise a window portion configured for transmission of
light, as may be useful in optical detection, optical
identification, or optical measurement of a sample or a portion or
constituent of a sample held in the vessel. A window portion may
include an optically flat surface, and may include two optically
flat surfaces. In embodiments, at least portions of two optically
flat surfaces may be substantially parallel to each other. A vessel
comprising two or more materials may comprise a flat area
configured for contact with a sonicator tip made from a first
material or a first combination of materials, and may comprise a
window, made from a second material or a second combination of
materials; the first material or combination of materials may be
different from the second material or combination of materials.
[0081] In embodiments, pathogen-identifying material may comprise a
nucleic acid, and may comprise a pathogen-identifying sequence of
nucleic acids. In embodiments, pathogen-identifying material may
comprise an amino acid, and may comprise a pathogen-identifying
sequence of amino acids. In embodiments, detecting the presence of
pathogen-identifying material comprises contacting said
pathogen-identifying material with a probe, which may be a labeled
probe. In embodiments, a probe, including a labeled probe, may
comprise nucleic acids. In embodiments, a probe, including a
labeled probe, may comprise a sequence of nucleic acids
complementary to at least a portion of said pathogen-identifying
material. In embodiments, a probe, including a labeled probe, may
comprise amino acids. In embodiments, a probe, including a labeled
probe, may comprise an antibody or an antibody fragment which
specifically binds to at least a portion of said
pathogen-identifying material.
[0082] In embodiments, a probe, or a complex, may comprise a label.
In embodiments, a label may be selected from the group consisting
of a dye, an epitope tag, a fluorescent moiety, a luminescent
moiety, a chemiluminescent moiety, an enzymatic label, a magnetic
label, a paramagnetic label, a contrast agent, a nanoparticle, a
radioisotope, biotin, streptavidin, and a quencher.
[0083] In embodiments, detecting the presence of
pathogen-identifying material may comprise detecting a complex
comprising pathogen-identifying material, or detecting a probe
bound to pathogen-identifying material in said sample. In
embodiments of the methods disclosed herein, a detecting step may
comprise detecting the presence of a label.
[0084] In embodiments, identifying a pathogen-identifying material
may comprise identifying a complex comprising pathogen-identifying
material, or detecting a probe bound to pathogen-identifying
material in said sample. In embodiments of the methods disclosed
herein, an identifying step may comprise detecting the presence of
a label.
[0085] In embodiments, measuring the amount of pathogen-identifying
material may comprise measuring the amount of a complex comprising
pathogen-identifying material, or measuring the amount of probe
bound to pathogen-identifying material in said sample. In
embodiments of the methods disclosed herein, a measuring step may
comprise measuring the amount of probe, or label, bound or
released, may comprise measuring a label or measuring the amount of
a label.
[0086] In embodiments, measuring the amount of pathogen-identifying
material may comprise a nucleic acid assay. In embodiments, a
nucleic acid assay for measuring the amount of pathogen-identifying
material may comprise a probe; for example, a probe may comprise
nucleic acids, or nucleic acid analogs, complementary to a target
nucleic acid, or complementary to a portion of a target nucleic
acid. In embodiments, measuring the amount of pathogen-identifying
material may comprise amplification of target nucleic acids, or may
comprise amplification of portions of target nucleic acids. In
embodiments, amplification of target nucleic acids, or
amplification of portions of target nucleic acids, may comprise
thermal cycling. In embodiments, amplification of target nucleic
acids, or amplification of portions of target nucleic acids, may
comprise isothermal methods of nucleic acid amplification.
[0087] In embodiments of the methods disclosed herein, detecting
probe bound to pathogen-identifying material may comprise an
optical measurement, identifying pathogen-identifying material may
comprise an optical measurement, and measuring the amount of probe
bound may comprise an optical measurement. In embodiments, optical
measurement may comprise detection of, or measurement of, the
intensity of electromagnetic radiation passing through, or emitted
from, a composition comprising said biological sample.
[0088] Methods, devices, and systems disclosed herein provide rapid
assays which require only small amounts of sample, such as only
small amounts of blood. Device and systems disclosed herein are
configured to perform such rapid assays which require only small
amounts of sample, such as only small amounts of blood, urine,
sputum, tears, material obtained from a nasal swab, throat swab,
cheek swab, or other biological sample. Accordingly, the methods,
devices, and systems provide rapid tests, which require only small
biological samples, and thus provide advantages over other methods,
compositions, assays, devices, and systems.
[0089] This Summary is provided to introduce a selection of
concepts in a simplified form that are further described below in
the Detailed Description. This Summary is not intended to identify
key features or essential features of the claimed subject matter,
nor is it intended to be used to limit the scope of the claimed
subject matter.
BRIEF DESCRIPTION OF THE DRAWINGS
[0090] FIG. 1 presents a schematic illustration of an embodiment of
a system as disclosed herein, including a sonicator having a
sonicator horn, a vessel configured to hold a sample, a vessel
holder configured to hold a vessel for sonication, a sample
handling system configured to place a sample within a vessel, a
detector for detecting pathogen-identifying material from a sample,
a communications assembly (labeled communication link/display),
which may comprise a display, for receiving instructions and
information, and for providing information and data to a user, or
to other components or equipment, a controller to operate the
components of the system, and a power supply to enable the
operation of the components.
[0091] FIG. 2 shows an example of a sonicator suitable for an
embodiment of the devices, systems, and methods disclosed herein.
In embodiments, a portion of the tip of a sonicator may touch or be
inserted into a sample solution in a vessel. In such embodiments,
the sonicator tip contacts the sample solution. In an embodiment of
configurations illustrated in this figure, the tip portion of the
sonicator may be prevented from contacting the sample by the
interposition of a compliant barrier material (e.g., the vessel may
have a compliant covering, such as a compliant plastic or rubber
covering). Pressing the sonicator tip onto a proximal side of the
compliant barrier, effective to cause a distal portion of the
compliant barrier to contact the solution within the vessel, allows
transfer of ultrasonic energy from the sonicator to the sample
solution in the vessel while preventing direct contact between the
sonicator tip and the sample solution.
[0092] FIG. 3 shows an example of a sonicator suitable for an
embodiment of the devices, systems, and methods disclosed herein,
in which a portion of the sonicator contacts a wall of the vessel
holding a sample solution; in such embodiments, no portion of the
sonicator contacts the sample solution, preventing contamination of
the sample and of the sonicator, so that there is no requirement to
clean or condition the sonicator tip before its re-use.
[0093] FIG. 4 shows an example of an embodiment of a sonicator
having a collet for holding a vessel. The collet may be cinched
around a vessel contained within the collet to provide a tight fit
between the collet and vessel; such a tight fit may aid in the
efficient transfer of ultrasonic energy between the transducer of
the sonicator (via the collet) and the vessel.
[0094] FIG. 5 shows an example of a sonicator mounted for use in an
embodiment of the devices disclosed herein. As shown, a distal
portion of the sonicator passes through a port in a vessel holder,
allowing direct contact between the sonicator and a wall of a
vessel held by the vessel holder. As shown, a vessel holder may
have a further aperture, allowing access to another portion of the
vessel, and including optical access without requiring an optical
path through a wall of the vessel holder. As shown, a vessel holder
may have walls without apertures; such walls may be configured for
optical access as well, by construction using suitable materials;
by providing a wall of proper flatness, orientation, or thickness
of the wall; by proper preparation or construction of the wall
surface, or by other means.
[0095] FIGS. 6A-6B provide views of an embodiment of a sonicator
and sonicator assembly suitable for devices, systems, and methods
as disclosed herein. FIG. 6A shows a side view of an embodiment of
a sonicator mounted with a solenoid configured to move the
sonicator horn in a transverse direction (e.g., towards the vessel
holder, or away from the vessel holder, as shown in the figure). A
spring is provided to urge the sonicator away from the vessel
holder upon release of solenoid force drawing the sonicator towards
the vessel holder. The sonicator horn is disposed so as to approach
a vessel held within the vessel holder. At rest (with the solenoid
off), as shown in FIG. 6A, the sonicator horn does not contact a
vessel held within the vessel holder, and the spring is in an
extended conformation.
[0096] FIG. 6B shows a side view of an embodiment of a sonicator
mounted with a solenoid configured to move the sonicator horn in a
transverse direction (e.g., towards the vessel holder, or away from
the vessel holder, as shown in the figure). A spring is provided to
urge the sonicator away from the vessel holder upon release of
solenoid force drawing the sonicator towards the vessel holder. The
sonicator horn is disposed so as to approach a vessel held within
the vessel holder. With the solenoid on, as shown in FIG. 6B, the
sonicator horn is in contact with an outer wall of a vessel held
within the vessel holder, and the spring is in a compressed
conformation. Operation of the sonicator in this configuration, in
which the sonicator horn is in contact with a wall of a vessel held
within the vessel holder, is effective to provide ultrasonic energy
to a sample solution within the vessel. Providing sufficient
ultrasonic energy for s sufficient amount of time is effective to
disrupt cells within the sample solution; in particular, pathogen
cells within the sample solution may be disrupted, effective to
release pathogen-identifying material for detection,
identification, and measurement.
[0097] FIG. 7 provides a further example of a solenoid assembly
suitable for use in the devices and systems disclosed herein. In
this figure, the vessel is covered with a cap; such a cap is
effective to prevent loss of sample fluid during sonication, and to
prevent spread of sample contents (by spillage, aerosolization, or
other means) outside the vessel. In this figure, a vessel is shown
held in a vessel holder, and a sonicator horn is shown in contact
with a side wall of the vessel. Such a configuration is an
operative configuration effective to disrupt cells within the
sample solution, such as pathogen cells within the sample
solution.
[0098] FIG. 8 provides a schematic illustration of a device having
a sonicator and other features as disclosed herein. The device
illustrated by the figure has a sonicator with a sonicator horn,
which can be made to contact a wall of a vessel (e.g., by motion of
a movable sonicator mount, as indicated in the figure). The vessel
can contain a sample solution, and is configured to be held by a
vessel holder. As shown, a vessel cap may be disposed on a surface
of the vessel. A sample handling system may be provided effective
to transport a biological sample, or a portion thereof, and
reagents if applicable, to the vessel. As discussed herein, a
sample handling system may also be configured to manipulate or move
a vessel, a vessel cap, or other article, within a housing of a
device. A device may have a detector, e.g., within the device
housing as illustrated in the figure, such as a detector configured
to detect pathogen-identifying materials released by sonication of
a sample solution. A device may further include other components
for performing assays, detecting analytes, and collecting and
communicating data obtained from biological samples.
[0099] FIG. 9 provides a schematic diagram of an embodiment of a
portion of a device having multiple, multiplexed sonicators. In a
device comprising multiple sonicators, such sonicators and the
associated multiplexer, power supply, cabling, controller, and
other associated components are typically placed within the housing
of a device. In a system comprising multiple sonicators, which may
not all be in the same device, such sonicators and the associated
multiplexer, power supply, cabling, controller, and other
associated components are typically placed within separate
housings, e.g., within the housings of the corresponding devices of
the system.
[0100] FIGS. 10A-D provide an example of embodiments of vessels
suitable for containing a sample solution for sonication. FIG. 10A
shows a side view of a vessel, with a flat wall surface facing
outward, and the opening (for filling the vessel) shown at the top.
The bottom of the vessel shown is also flat; the flat side wall and
the flat bottom are configured to contact a tip portion of a
sonicator, such as a tip of a sonicator horn. The wider portion at
the top of the vessel includes surfaces and mating sockets. The
mating sockets comprise recesses configured for engagement of a
transport and/or force-providing member (e.g., a nozzle of a sample
handling system) which a) allows transport of the vessel and b)
provides a surface for provision of downward force to oppose upward
force of a sonicator horn placed on a flat bottom surface of the
vessel. FIG. 10B shows a side view of a vessel as shown in FIG.
10A, with a flat wall surface facing rightward, and the opening
(for filling the vessel) shown at the top. FIG. 10C shows a vessel
as shown in FIGS. 10A and 10B, with a flat wall surface facing
leftward and slightly upward, and the opening (for filling the
vessel) shown at the top. Openings of the mating sockets are
visible in this view, as is an inner ridge within the internal
chamber of the vessel. An inner ridge as illustrated in the figure
may be configured to seat, or to seal, a cap. FIG. 10D shows a
cross-sectional view of a vessel as shown in FIGS. 10A-C, with the
opening (for filling the vessel) shown at the top. The ridge and
inner chamber of the vessel, and the shape and depth of the mating
sockets are visible in this cross-sectional view.
[0101] FIGS. 11A-D provide further embodiments of vessels suitable
for containing a sample solution for sonication. FIG. 11A shows an
embodiment of a tubular vessel with a rounded bottom. FIG. 11B
shows an embodiment of a conical vessel with a rounded bottom. FIG.
11C an embodiment of an elongated tubular vessel with a rounded
bottom. FIG. 11D shows an embodiment of a wide tubular vessel with
a rounded bottom.
[0102] FIGS. 12A-D provide yet further embodiments of vessels
suitable for containing a sample solution for sonication. FIG. 12A
shows an embodiment of a tubular vessel with a rounded bottom and a
tapered (e.g., partially conical-shaped cap). FIG. 12B shows an
embodiment of a conical vessel with a rounded bottom having flat
side surfaces configured to engage with a sonicator tip. FIG. 12C
an embodiment of an elongated tubular vessel with a rounded bottom
and a protruding flat surface configured to engage with a sonicator
tip. FIG. 12D shows an embodiment of a wide tubular vessel with a
rounded bottom having a cap connected to the vessel via cap
linkage.
[0103] FIG. 13 illustrates a configuration in which a vessel having
a flat bottom is disposed to contact a tip of a sonicator. Such
contact is effective to transfer ultrasonic energy to the wall of
the vessel and thereby to a sample solution contained within the
vessel, effective to disrupt cells in the sample solution when the
sonicator is activated. Functional contact between the vessel and
the sonicator may be aided or improved by the spring shown in the
figure. (Note that the action and function of the spring in this
figure is opposite to that of the springs shown in FIGS. 6A, 6B,
and 7. The present spring acts to urge the sonicator and vessel
together; the springs shown in FIGS. 6A, 6B, and 7 act to urge the
sonicator and vessel apart from each other.) Functional contact
between the vessel and the sonicator may be aided or improved by
application of force (as illustrated by the downward arrow in the
figure) urging the vessel onto the tip of the sonicator horn,
improving functional contact and improving energy transfer from the
sonicator to the sample solution and cells within it.
[0104] FIG. 14 presents the results of quantitative polymerase
chain reaction (qPCR) measurements made from pathogen-identifying
material released for assay by sonication according to the present
methods. The horizontal axis represents the number of cycles while
the vertical axis (in relative light units) represents the numbers
of copies of the target nucleic acid sequence indicative of the
target pathogen. As shown in the figure, all target nucleic acids
were detectable after about 25 cycles.
DETAILED DESCRIPTION
[0105] Applicants provide devices, systems, and methods for
disrupting cells using a sonicator. Sonicators provide ultrasonic
energy which disrupts cells by producing, for example, cavitation
within a sample fluid containing the cells, which cells may be
disrupted by sonoporation due to cavitation in the fluid. In
embodiments, such devices, systems, and methods for disrupting
cells using a sonicator may be applied to an external surface of a
vessel, where the vessel contains a biological sample suspected of
harboring a pathogen or multiple pathogens. A sonicator may provide
ultrasonic energy to a vessel, for example, by contacting a wall of
the vessel with its tip portion (e.g., with a tip of a sonicator
horn), effective to transfer energy through the vessel wall to
fluid within the vessel. Such fluid may comprise a biological
sample. Applicants further disclose vessels for use with sonicators
as disclosed herein and with devices and systems comprising such
sonicators.
[0106] Accordingly, embodiments of devices and systems for
detecting, identifying, or measuring pathogen-identifying material
in at least a portion of a biological sample, e.g., of blood,
urine, sputum, material obtained from a nasal swab, a throat swab,
a cheek swab, or other sample; and embodiments of devices and
systems for detecting, identifying, or measuring
pathogen-identifying material in at least a portion of a biological
sample, e.g., of blood, urine, sputum, material obtained from a
nasal swab, a throat swab, a cheek swab, or other sample, and at
least one other biologically relevant attribute from said
biological sample, e.g., of blood, urine, sputum, material obtained
from a nasal swab, a throat swab, a cheek swab, or other sample,
from a subject are disclosed herein.
[0107] Devices, systems, and methods disclosed herein may comprise,
and may be used with, devices, systems, and methods as disclosed
in, for example, U.S. Pat. No. 8,088,593; U.S. Pat. No. 8,380,541;
U.S. Pat. App. Ser. No. 61/799,533, filed Mar. 15, 2013; U.S.
patent application Ser. No. 13/769,798, filed Feb. 18, 2013; U.S.
patent application Ser. No. 13/769,779, filed Feb. 18, 2013; U.S.
patent application Ser. No. 13/244,947 filed Sep. 26, 2011;
PCT/US2012/57155, filed Sep. 25, 2012; U.S. application Ser. No.
13/244,946, filed Sep. 26, 2011; U.S. patent application Ser. No.
13/244,949, filed Sep. 26, 2011; and U.S. Application Ser. No.
61/673,245, filed Sep. 26, 2011, the disclosures of which patents
and patent applications are all hereby incorporated by reference in
their entireties.
[0108] Devices disclosed herein comprise a sonicator configured to
transfer ultrasonic energy to a sample solution. In embodiments, a
sonicator, e.g., a sonicator tip, may contact a sample solution
directly. In embodiments, a sonicator, e.g., a sonicator tip, may
contact a sample solution indirectly via a barrier material. In
embodiments, a barrier material may be a compliant material. In
embodiments, a sonicator, e.g., a sonicator tip, may not contact a
sample solution directly, but may instead be configured to contact
a wall or other surface of a vessel containing a solution to which
ultrasonic energy is to be directed.
[0109] A sonicator may comprise a sonicator horn, e.g., may
comprise an elongated portion configured to transmit and/or
concentrate ultrasonic energy at a small area distal from the
oscillator of the sonicator. A sonicator (and its sonicator horn)
may have a tip configured to contact and to interact with a vessel
containing a solution to which ultrasonic energy is to be directed.
In embodiments, a sonicator may be mounted on a sonicator mount,
which may be configured to hold a sonicator in an operating
position during use. In embodiments, a sonicator may be mounted on
a sonicator mount, which may be configured to place a sonicator in
a disengaged position when not in use transmitting ultrasonic
energy.
[0110] In embodiments, a sonicator mount may provide for movement
of the sonicator, e.g., may allow the engagement and disengagement
of the sonicator with a vessel wall, as needed for the operation of
the sonicator. For example, a device comprising a sonicator may
include a sonicator mount operably connected with a solenoid
configured to place a sonicator tip in contact with a vessel
containing a sample solution when the solenoid is activated, where
the sonicator is mounted effective that the sonicator tip is
removed from contact with the vessel when the solenoid is not
activated. In embodiments, a spring may be used to aid in removing
a sonicator tip from contact with a vessel when a solenoid is not
activated.
[0111] In embodiments, a sample solution may be contained in a
vessel, and a portion of a sonicator may be applied to an external
wall of the vessel. In embodiments, the external wall to which a
sonicator is applied may be a side wall of the vessel. In
embodiments, the external wall to which a sonicator is applied may
be a bottom wall of the vessel. In embodiments, the external wall
to which a sonicator may be applied is a top wall of the vessel. In
embodiments, a sonicator may be applied to more than one wall of
the vessel.
[0112] A vessel comprises an internal chamber configured for
holding fluid, such as a sample solution (although it will be
understood that such an internal chamber may hold a reagent, or
water, or any other fluid). A vessel will typically include an
opening configured to accept a sample solution; for example, a
vessel may be filled (partially or completely) via an opening.
Where the vessel has only a single opening leading to an inner
chamber, that opening defines the top of the vessel. In
embodiments, the vessel is open to the environment, e.g., has an
open passage at its top. In embodiments, a vessel may be configured
to accept a cap, where the cap is effective to occlude an opening
in the vessel, e.g., to close an open passage at the top of the
vessel. In embodiments, a cap may be a separate element (e.g., may
be able to be completely separated from the vessel); in
embodiments, a cap may be a portion of a vessel capable of moving
into and out of a position that occludes an opening or passage that
(when not occluded) provides access to an interior portion of the
vessel. In embodiments, a cap may comprise a portion of a vessel.
For example, a cap may be connected to other portions of a vessel
by a hinge, or by a thread, or cable, or other flexible connector
of any kind. In embodiments, the vessel is closed, and not open to
the environment, when ultrasonic energy is applied to it. A cap,
when in place occluding an opening of a vessel, may be effective to
prevent or reduce spilling of a sample solution held within the
vessel. A cap, when in place occluding an opening of a vessel, may
be effective to prevent or reduce loss of sample solution due to
splashing, bubble formation, aerosolization, or other actions due
to delivery of ultrasonic energy to a sample solution held within
the vessel.
[0113] In embodiments of the devices, systems and methods disclosed
herein, a sample handling system may be used to transport and
deliver a sample solution to a vessel, and to fill a vessel (either
partially or fully) with a sample solution. In embodiments, a
sample handling system comprises a pipette. A pipette may be
configured to accept a pipette tip, e.g., to mount and transport a
pipette tip attached to the pipette. In embodiments, a pipette
comprises a nozzle configured to accept a pipette tip. A pipette
may be configured to aspirate a fluid, such as a sample solution,
into a pipette tip attached to the pipette (e.g., a pipette tip
which is attached to a nozzle of the pipette). In embodiments, a
pipette may be configured to dispense a fluid, such as a sample
solution, from a pipette tip attached to the pipette (e.g., to a
nozzle of the pipette). A pipette may be configured to transmit
force to a surface or component of a device. In embodiments, a
pipette nozzle may contact a surface or component of a device,
effective to transmit force to that surface or component. In
embodiments, a pipette nozzle may contact a mating recess of a
vessel, and, in embodiments, may engage a mating recess of a
vessel. In embodiments, two, or more pipette nozzles may contact
mating recesses of a vessel, and, in embodiments, may engage mating
recesses of a vessel. In embodiments, a pipette of a sample
handling system may be movable, and is preferably movable in at
least two, and more preferably in three dimensions (e.g., is
movable in one, two, or all three of horizontally, laterally, and
vertically).
[0114] In embodiments, devices comprising a sonicator, and systems
and methods comprising or using such devices, may comprise a
detector configured to detect pathogen-identifying material in a
sample. A detector may be, for example, an optical detector, such
as a spectrophotometer, a photomultiplier, a charge-coupled device,
a camera, or other device or system configured to detect a
light-based signal indicative of the presence of a
pathogen-identifying material. In embodiments, a detector may be
configured to, or be effective to, detect a signal comprising
chemiluminescence, luminescence, fluorescence, absorbance,
transmittance, turbidity, a color change, or other change in light,
whether emitted, transmitted, or absorbed, effective to signal the
presence of a pathogen-identifying material in a sample. In
embodiments, a detector may comprise an electrochemical detector,
or a temperature sensor, or a pH sensor, or a radiation sensor, or
an ion-sensitive electrode, or other sensor capable of detecting
the presence of a pathogen-identifying material in a sample.
[0115] Methods for detecting pathogen-identifying material include
assays for detecting nucleic acids (e.g., DNA or RNA), assays for
detecting peptides and proteins (including glycoproteins), assays
for detecting other pathogen-related molecules, complement fixation
assays, hemagglutination assays (e.g., for influenza), and other
assays. Methods for detecting nucleic acids include polymerase
chain reaction (PCR) methods (see, e.g. U.S. Pat. No. 4,683,202).
PCR methods include quantitative PCR (qPCR), reverse-transcriptase
PCD (RT-PCR), "real-time" PCR, one-step PCR, two-step PCR, and
other methods known in the art. Methods for detecting nucleic
acids, including methods for nucleic acid amplification, may
include thermal cycling, as in PCR methods for example. However,
amplification of target nucleic acids, or amplification of portions
of target nucleic acids, may comprise isothermal methods of nucleic
acid amplification. Isothermal nucleic acid amplification methods
include, for example, loop-mediated isothermal amplification
("LAMP") (see, e.g. U.S. Pat. No. 6,410,278), and other isothermal
methods. An exemplary isothermal nucleic acid amplification method
suitable for use with the devices, systems, methods, and vessels
disclosed herein are disclosed, for example, in U.S. Patent
Application 61/800,606, filed Mar. 15, 2013, the entire contents of
which application is hereby incorporated by reference in its
entirety.
[0116] Methods for detecting peptides and proteins include enzyme
immunoassays such as Enzyme-Linked ImmunoSorbent Assays (ELISAs)
and other assays utilizing antibodies or antibody fragments,
complement-based reactions, measurement of absorbance of
ultraviolet or other frequency of light, assays utilizing specific
receptor-ligand interactions, and other assays known in the art.
Assays for detecting other pathogen-related molecules include
assays for bacterial sugars and lipids (e.g., bacterial
lipopolysaccharide (LPS)), and other assays known in the art. A
detector for use with such assays may be an optical detector, a pH
detector, an electrochemical detector, a temperature sensor, an
ion-sensitive electrode, a radiation detector, or other
detector.
[0117] In embodiments, devices comprising a sonicator, and systems
and methods comprising or using such devices, may comprise a
controller. In embodiments, a controller may comprise a processor.
In embodiments, a controller may be connected to, and may control
the operation of, components of a device; such components are
typically disposed within a housing of the device. In embodiments,
a controller may control the operation of a sonicator. In
embodiments, a controller may control the operation of a sample
handling system. In embodiments, a controller may control the
operation of a detector. In embodiments, a controller may control
the operation of any component or unit of the device. Other
components may include, for example, a camera, a chemistry assay
unit, a nucleic acid assay unit, a heating unit, a communication
unit, a protein chemistry unit, or other component or unit. In
embodiments, a controller may control the operation of one or more
components of a device according to a protocol. In embodiments, a
protocol by which a controller controls the operation of any one or
more component or unit of a device may be preprogrammed, e.g., may
be resident on the device. In embodiments, a protocol by which a
controller controls the operation of any one or more component or
unit of a device may be obtained from another device, or from a
user, or from a laboratory, or from a network, or from the cloud.
In embodiments, a protocol by which a controller controls the
operation of any one or more component or unit of a device may be
updated, or may be updatable, according to information or
instructions from another device, or from a user, or from a
laboratory, or from a network, or from the cloud. In embodiments, a
device may receive information, or instructions, or updates, or
protocols, via a user interface. In embodiments, a device may
receive information, or instructions, or updates, or protocols, via
a communication assembly.
[0118] In embodiments, devices comprising a sonicator, and systems
and methods comprising or using such devices, may comprise a
display effective to provide a user with information regarding the
operation of the device, information regarding the progress of an
assay performed by the device, or information regarding the results
of an assay performed by the device. In embodiments, a display may
comprise a visual display, or may comprise a printed display, or
may comprise an audio signal, which may include an audio signal
understandable as speech by a user, or may comprise any combination
or all of such displays. In embodiments, a display may comprise a
user interface. In embodiments in which a display comprises a user
interface, a device may receive, e.g., information, commands,
protocols, or other input.
[0119] In embodiments, devices comprising a sonicator, and systems
and methods comprising or using such devices, may comprise a
communication assembly effective to communicate with one or more of
a user, another device, a laboratory, a network, the cloud, or
other communication target. In embodiments, a communication
assembly may provide a communication target with information
regarding the operation of the device, information regarding the
progress of an assay performed by the device, or information
regarding the results of an assay performed by the device. In
embodiments, a communication assembly may be configured to allow a
device to receive, e.g., information, commands, protocols, or other
input from an outside source, such as, e.g., a user, another
device, a laboratory, a network, the cloud, or other communication
source.
Definitions
[0120] Before the present devices, systems, and methods are
disclosed and described, it is to be understood that the
terminology used herein is for the purpose of describing particular
embodiments only and is not intended to be limiting. It is also to
be understood that the present disclosure provides explanatory and
exemplary descriptions and examples, so that, unless otherwise
indicated, the devices, systems, and methods disclosed herein are
not limited to the specific embodiments described herein.
[0121] It must be noted that, as used in the specification and the
appended claims, the singular forms "a," "an" and "the" include
plural referents unless the context clearly dictates otherwise.
Thus, for example, reference to "a salt" refers to a single salt or
mixtures of different salts, and the like.
[0122] In this specification and in the claims that follow,
reference will be made to a number of terms, which shall be defined
to have the following meanings:
[0123] Acronyms and abbreviations, such as "rpm" (revolutions per
minute), "min" (minute), "sec" (second), and so forth, have their
customary meanings.
[0124] As used herein, the term "assay" and grammatical equivalents
refers to tests, measurements, observations, and other experimental
procedures which may be applied to a sample for detection of an
analyte, identification of an analyte, and measurement of the
amounts of an analyte in a sample. Assays may be physical assays
which detect, identify, or measure a physical property of a sample;
assays may be chemical assays, which detect, identify, or measure a
chemical property of a sample, or perform chemical reactions in or
with a sample; and include assays which use optical, electrical or
electronic, chemical, or other means of detection and
measurement.
[0125] As used herein the term "ultrasound" refers to vibrations at
frequencies beyond those capable of being detected by a human
subject, i.e., frequencies greater than about 20,000 cycles per
second (20 kiloHerz (kHz)).
[0126] As used herein the term "ultrasonic energy" refers to the
energy (e.g., mechanical energy) of vibrations at frequencies
beyond those capable of being detected by a human subject, i.e.,
frequencies greater than about 20,000 cycles per second (20 kHz). A
sonicator may be effective to produce and apply ultrasonic
energy.
[0127] As used herein, the term "sonicator" refers to a device
effective to provide ultrasonic energy, e.g., by providing
mechanical energy, typically in the form of vibrations, at
ultrasonic frequencies. A sonicator may include a driving element
which provides high-frequency vibrations; for example, many
sonicators utilize piezoelectric materials to produce
high-frequency vibrations. Piezoelectric ultrasound generators
typically require high voltages (e.g., provided by a time-varying
electrical input having peak voltages of about 200 volts (V) to
about 400 V, e.g., about 50 root-mean-square volts (VRMS) to about
300 VRMS) to provide the needed alternating current drive to
operate such generators.
[0128] As sonicator has a tip portion, which includes the distal
tip and portions nearby; ultrasonic energy from a sonicator is
typically transmitted to a target material by the tip portion.
Sonicators often have sonicator horns, often made from stainless
steel, for example, configured to direct ultrasonic energy to a tip
portion; where a sonicator has a sonicator horn, the tip of the
sonicator horn comprises the sonicator tip, and the tip portion of
the sonicator horn comprises the sonicator tip portion.
[0129] As used herein, the terms "horn" and "sonicator horn" refer
to a portion of a sonicator configured to focus ultrasonic energy,
or to provide a pathway for the direction of ultrasonic energy, to
a destination. A horn may be designed and configured to transfer or
transmit ultrasonic energy in an efficient manner, and may, for
example, be configured to resonate at particular frequencies which
may be tuned or adapted to particular frequencies of choice of a
driving element of a sonicator. A horn may have a tip, or tip
portion; a horn tip or tip portion may be configured to maximize
energy transfer, e.g., by maximizing contact, to a vessel, or wall
of a vessel, or to a fluid contained within a vessel.
[0130] As used herein, the terms "fill" and "filled" and their
grammatical equivalents, e.g., as used in phrases such as "a vessel
may be filled with a sample solution" refer to the transfer of any
amount, including partial filling and complete filling. These terms
as used herein do not require that such filling completely fill a
container, but include any lesser amount of filling as well.
[0131] As used herein, the term "translucent" means that light (or
other optical signal) is able to pass through; thus, a translucent
material will allow light or other optical signals to pass through
the material.
[0132] As used herein, the term "transparent" means that light (or
other optical signal) is able to pass through with little or no
distortion; thus, a transparent material will allow light or other
optical signals to pass through the material without causing
significant distortion of the signal, and thus allows imaging
through the material with little or no distortion.
[0133] A vessel may be made with, a glass; silicon; plastic;
ceramic; polymer; or other material or combination of materials. A
glass may be, or may include, for example, sodium silicate,
borosilicate, aluminosilicate, quartz, or other glass. A polymer,
or polymeric material, may be or may include, for example,
polypropylene, polycarbonate, polystyrene, polyurethane,
polyethylene, polyacrylamide, polyacrylate, polymethacrylate,
polymethylmethacrylate (PMMA), poly(4-methylbutene), other acrylic,
polydimethysiloxanes (PDMS), polyvinylchloride (PVC), poly(vinyl
butyrate) polysulfone, acrylonitrile-butadiene-styrene (ABS),
poly(ethylene terephthalate), a fluorocarbon polymer (e.g.,
polytetrafluoroethylene (PTFE or Teflon.RTM.)), nylon, a
co-polymer, or combinations thereof. A ceramic may be, or may
include, for example, silicon carbide, tungsten carbide, aluminum
oxide, and others.
[0134] As used herein, the terms "detect", "detection", "detecting"
and grammatical equivalents refer to a determination that a target
analyte, such as a pathogen-identifying material, is present.
Detection does not require that a minimum amount of the target
analyte be present in the sample, but merely that the analyte be
observed to be present, whether directly or indirectly (although in
practical terms, a lower limit below which detection is unlikely to
occur may exist).
[0135] As used herein, the terms "identify", "identification",
"identifying", and grammatical equivalents refer to a determination
of the identity of a target analyte found in a sample. Such
identification, in addition to detection, may be useful where
detection may be ambiguous as to the specific identity of a
detected analyte; for example, where an influenza virus is
detected, it may further be useful to identify the particular
strain, or strains, of influenza found in the sample. Thus,
identification may provide more detailed and specific information
than does detection.
[0136] As used herein, the terms "measure", "measurement",
"measuring", and grammatical equivalents refer to a determination
of the amounts of a target analyte found in a sample. Thus,
measurement provides quantitative information which may not be
provided by detection or identification, or which may be more
precise that information provided by detection or
identification.
[0137] The term "nucleic acid" refers to nucleotides and
nucleosides which make up, for example, deoxyribonucleic acid (DNA)
macromolecules and ribonucleic acid (RNA) macromolecules. Nucleic
acids may be identified by the base attached to the sugar (e.g.,
deoxyribose or ribose); nucleic acid sequences may be used to
identify an organism from which the nucleic acid sequence was
obtained.
[0138] The terms "polypeptide" and "protein" may be used
interchangeably to refer to molecules comprised of amino acids
linked by peptide bonds. Individual amino acids may be termed
"residues" of a polypeptide or protein. Unique amino acid sequences
of polypeptides may be used to identify the organisms from which
the polypeptides were obtained.
[0139] The term "antibody" is used in the broadest sense and
specifically covers single monoclonal antibodies (including agonist
and antagonist antibodies), antibody compositions with polyepitopic
specificity, and antibody fragments. Monoclonal antibodies are
obtained from a population of substantially homogeneous antibodies,
i.e., the individual antibodies comprising the population are
identical except for possible naturally-occurring mutations that
may be present in minor amounts. For example, monoclonal antibodies
may be made by the hybridoma method first described by Kohler et
al., Nature, 256:495 (1975), or may be made by recombinant DNA
methods (see, e.g., U.S. Pat. No. 4,816,567 to Cabilly et al.).
[0140] "Antibody fragment", and all grammatical variants thereof,
as used herein is defined as a (1) portion of an intact antibody
comprising the antigen binding site or variable region of the
intact antibody, wherein the portion is free of the constant heavy
chain domains (i.e. CH2, CH3, and CH4, depending on antibody
isotype) of the Fc region of the intact antibody, and (2)
constructs comprising a portion of an intact antibody (as defined
by the amino acid sequence of the intact antibody) comprising the
antigen binding site or variable region of the intact antibody.
Examples of antibody fragments include Fab, Fab', Fab'-SH, F(ab')2,
Fd, Fc, Fv, diabodies, and any other "Non-single-chain
antigen-binding unit" as described, e.g., in U.S. Pat. No.
7,429,652.
[0141] The term "pathogen" as used herein refers to an agent that
may cause a disease, such as an infectious disease, in a subject.
Pathogens include, for example, Gram-negative bacteria,
Gram-positive bacteria, other bacteria, RNA viruses, DNA viruses,
prions, yeast, fungi, protozoans, helminths, nematodes, and any
other pathogenic agent which may sicken a subject or, if
transmitted from a subject who may not suffer disease, could cause
disease in a further subject to which the pathogen is
transmitted.
[0142] As used herein, "pathogen-identifying material" refers to
proteins and nucleic acids derived from a pathogen, such as a
bacterium, a prion, a virus, a yeast cell, a fungus, a protozoan
(also termed "protist"), a helminth, nematode, or other viral,
single cellular or multicellular organism which may be found in a
biological sample collected from a subject. Pathogen-identifying
material may comprise a target nucleic acid. Pathogen-identifying
material may comprise a target peptide, or epitope (e.g., a portion
of a peptide). The proteins, and the nuclear material of such
organisms (whether DNA or RNA), is indicative of the particular
type of organism, and may be used to detect the presence of such an
organism, and to identify it. Quantitative measurements of
pathogen-identifying material provide measurements of the amounts
of such organisms in a biological sample, and may be used to
determine the severity of an infection, or to track the progress of
a disease, and to track the course and success of its
treatment.
[0143] The term "probe" as used herein refers to a material which
is useful to identify a target compound or cell; a probe may be,
e.g., a nucleic acid probe, a protein probe, or other probe. A
target compound may be produced by, or found in or on, a prion, a
virus, a bacterium, a yeast, a fungus, a protozoan, or other
pathogen. A probe, as used herein, may comprise RNA or DNA, or
analogs of RNA or DNA, effective that the probe may recognize and
bind (e.g., hybridize) to a target nucleic acid. A target nucleic
acid may be, e.g., a viral, bacterial, yeast, fungal, protozoan,
helminth, nematode, or other nucleic acid sequence (which may be a
portion of a longer nucleic acid sequence). A probe, as used
herein, may comprise a polypeptide or protein, such as an antibody
or antibody fragment, which may recognize and bind to a target
compound or portion of a compound (e.g., an epitope of a protein
that is indicative of a target pathogen).
[0144] As used herein, a "marker", a "label", a "label" and a
"label moiety" refer to a detectable compound or composition which
is conjugated directly or indirectly to the antibody so as to
generate a "labeled" antibody. A label or marker provides a
detectable signal for at least the time period during which a
signal is to be observed. The label or marker may be detectable by
itself (e.g. radioisotope labels or fluorescent labels) or, in the
case of an enzymatic label, may catalyze chemical alteration of a
substrate compound or composition which is detectable.
[0145] A marker or label may be linked (non-covalently or
covalently) to a nucleic acid probe, an antibody or antibody
fragment, or other probe. A label may alter its detectability
(e.g., become detectable, or increase, or decrease its
detectability) upon contact with its target. A label may detach
from its probe upon contact between the probe and its target, and
thus may alter its detectability. Such alterations in detectability
are useful in assays for detecting, identifying, or measuring
analytes in a sample.
[0146] A label may be, for example, a dye, an epitope tag, a
fluorescent moiety, a luminescent moiety, a chemiluminescent
moiety, an enzymatic label, a magnetic label, a paramagnetic label,
a contrast agent, a nanoparticle, a radioisotope, biotin,
streptavidin, and a quencher. A nanoparticle may be a particle of
an element, such as a gold nanoparticle, or of an alloy or
compound, such as a quantum dot (a particle of a semiconductor
material), or other particle having a size typically in a range
between about 1 nm to about 100 nm.
[0147] As used herein, the terms "sample" and "biological sample"
refer to a blood, urine, sputum, tears, material from a nasal swab,
throat swab, cheek swab, or other bodily fluid, excretion,
secretion, or tissue obtained from a subject. These terms are
inclusive of an entire sample and of a portion of a sample. As used
herein, reference to a fluid sample includes reference to a sample
and a biological sample. Such samples may include fluids into which
material has been deposited, where such material may be obtained
from a nasal swab, throat swab, cheek swab, or other sample which
may include solid or semi-solid material, whether along with or
without natural fluids. Such fluids and samples comprise fluid
samples and sample solutions.
[0148] As used herein the "sample solution" refers to a fluid
sample itself, and to dilutions, mixtures, aliquots, or other
solutions which contain at least a portion of a sample.
[0149] As used herein the terms "sample handling system", "fluid
handling system" and grammatical equivalents refer to systems
configured to obtain, transport, and deliver fluids. In embodiments
disclosed herein, such systems comprise pipettes, nozzles, pipette
tips, mechanical components configured to move a pipette, a nozzle,
or a pipette tip to a desired location. Such a desired location is
typically within a housing of a device. In embodiments, a pipette
tip may be mounted on a nozzle; in embodiments, a pipette tip may
be removably mounted on a nozzle, effective that a nozzle may
engage and mount a first pipette tip, use the first pipette tip,
discard the first pipette tip, and then engage and mount a second
pipette tip. Such systems comprise means for aspirating liquid into
a pipette tip. Such systems comprise means for dispensing liquid
from a pipette tip. In embodiments of such systems, a pipette and
nozzle may engage and mount an element other than a pipette tip;
for example, in embodiments disclosed herein, a pipette and nozzle
may engage and mate with a mating socket of a vessel (see, e.g.,
FIGS. 10 and 11). In embodiments, a pipette and nozzle mated with a
mating socket of a vessel may be used to transport the vessel to a
desired location within a device. In embodiments, a pipette and
nozzle mated with a mating socket of a vessel may be used to apply
force to a vessel (see, e.g., FIG. 11 for a configuration where
such application of force may be useful).
[0150] The methods disclosed herein can be readily incorporated
into and used in device for processing a sample, or a system for
processing a sample, which may be an automated assay device, or may
be an automated assay system. Such assay devices and assay systems
may comprise devices and systems disclosed, for example, in U.S.
Pat. No. 8,088,593; U.S. Pat. No. 8,380,541; U.S. patent
application Ser. No. 13/769,798, filed Feb. 18, 2013; U.S. patent
application Ser. No. 13/769,779, filed Feb. 18, 2013; U.S. patent
application Ser. No. 13/244,947 filed Sep. 26, 2011;
PCT/US2012/57155, filed Sep. 25, 2012; U.S. application Ser. No.
13/244,946, filed Sep. 26, 2011; U.S. patent application Ser. No.
13/244,949, filed Sep. 26, 2011; and U.S. Application Ser. No.
61/673,245, filed Sep. 26, 2011, the disclosures of which patents
and patent applications are all hereby incorporated by reference in
their entireties.
[0151] Such a device, and such a system, may be useful for the
practice of the methods disclosed herein. For example, a device may
be useful for receiving a sample. A device may be useful for
preparing, or for processing a sample. A device may be useful for
performing an assay on a sample. A device may be useful for
obtaining data from a sample. A device may be useful for
transmitting data obtained from a sample. A device may be useful
for disposing of a sample following processing or assaying of a
sample.
[0152] A device may be part of a system, a component of which may
be a sample processing device. A device may be a sample processing
device. A sample processing device may be configured to facilitate
collection of a sample, prepare a sample for a clinical test, or
effect a chemical reaction with one or more reagents or other
chemical or physical processing, as disclosed herein. A sample
processing device may be configured to obtain data from a sample. A
sample processing device may be configured to transmit data
obtained from a sample. A sample processing device may be
configured to analyze data from a sample. A sample processing
device may be configured to communicate with another device, or a
laboratory, or an individual affiliated with a laboratory, to
analyze data obtained from a sample.
[0153] A sample processing device may be configured to be placed in
or on a subject. A sample processing device may be configured to
accept a sample from a subject, either directly or indirectly. A
sample may be, for example, a biological sample, e.g., of blood,
urine, sputum, material obtained from a nasal swab, a throat swab,
a cheek swab, or other sample, (e.g., a sample obtained from a
fingerstick, or from venipuncture, or an arterial biological
sample, e.g., of blood, urine, sputum, material obtained from a
nasal swab, a throat swab, a cheek swab, or other sample,), a urine
sample, a biopsy sample, a tissue slice, stool sample, or other
biological sample; a water sample, a soil sample, a food sample, an
air sample; or other sample. A biological sample, e.g., of blood,
urine, sputum, material obtained from a nasal swab, a throat swab,
a cheek swab, or other sample, may comprise, e.g., whole blood,
plasma, or serum. A sample processing device may receive a sample
from the subject through a housing of the device. The sample
collection may occur at a sample collection site, or elsewhere. The
sample may be provided to the device at a sample collection
site.
[0154] In some embodiments, a sample processing device may be
configured to accept or hold a cartridge. In some embodiments, a
sample processing device may comprise a cartridge. The cartridge
may be removable from the sample processing device. In some
embodiments, a sample may be provided to the cartridge of the
sample processing device. Alternatively, a sample may be provided
to another portion of a sample processing device. The cartridge
and/or device may comprise a sample collection unit that may be
configured to accept a sample.
[0155] A cartridge may include a sample, and may include reagents
for use in processing or testing a sample, disposables for use in
processing or testing a sample, or other materials. Following
placement of a cartridge on, or insertion of a cartridge into, a
sample processing device, one or more components of the cartridge
may be brought into fluid communication with other components of
the sample processing device. For example, if a sample is collected
at a cartridge, the sample may be transferred to other portions of
the sample processing device. Similarly, if one or more reagents
are provided on a cartridge, the reagents may be transferred to
other portions of the sample processing device, or other components
of the sample processing device may be brought to the reagents. In
some embodiments, the reagents or components of a cartridge may
remain on-board the cartridge. In some embodiments, no fluidics are
included that require tubing or that require maintenance (e.g.,
manual or automated maintenance).
[0156] A sample or reagent may be transferred to a device, such as
a sample processing device. A sample or reagent may be transferred
within a device. Such transfer of sample or reagent may be
accomplished without providing a continuous fluid pathway from
cartridge to device. Such transfer of sample or reagent may be
accomplished without providing a continuous fluid pathway within a
device. In embodiments, such transfer of sample or reagent may be
accomplished by a sample handling system (e.g., a pipette); for
example, a sample, reagent, or aliquot thereof may be aspirated
into an open-tipped transfer component, such as a pipette tip,
which may be operably connected to a sample handling system which
transfers the tip, with the sample, reagent, or aliquot thereof
contained within the tip, to a location on or within the sample
processing device. The sample, reagent, or aliquot thereof can be
deposited at a location on or within the sample processing device.
Sample and reagent, or multiple reagents, may be mixed using a
sample handling system in a similar manner. One or more components
of the cartridge may be transferred in an automated fashion to
other portions of the sample processing device, and vice versa.
[0157] A device, such as a sample processing device, may have a
sample handling system (also termed herein a fluid handling
system). A sample handling system may perform, or may aid in
performing, transport, dilution, extraction, aliquotting, mixing,
and other actions with a fluid, such as a sample. In some
embodiments, a sample handling system may be contained within a
device housing. A sample handling system may permit the collection,
delivery, processing and/or transport of a fluid, dissolution of
dry reagents, mixing of liquid and/or dry reagents with a liquid,
as well as collection, delivery, processing and/or transport of
non-fluidic components, samples, or materials. The fluid may be a
sample, a reagent, diluent, wash, dye, or any other fluid that may
be used by the device, and may include, but not limited to,
homogenous fluids, different liquids, emulsions, suspensions, and
other fluids. A sample handling system, including without
limitation a pipette, may also be used to transport vessels (with
or without fluid contained therein) around the device. The sample
handling system may dispense or aspirate a fluid. The sample may
include one or more particulate or solid matter floating within a
fluid.
[0158] In embodiments, a sample handling system may comprise a
pipette, pipette tip, syringe, capillary, or other component. The
sample handling system may have portion with an interior surface
and an exterior surface and an open end. The sample handling system
may comprise a pipette, which may include a pipette body and a
pipette nozzle, and may comprise a pipette tip. A pipette tip may
or may not be removable from a pipette nozzle. In embodiments, a
sample handling system may use a pipette mated with a pipette tip;
a pipette tip may be disposable. A tip may form a fluid-tight seal
when mated with a pipette. A pipette tip may be used once, twice,
or more times. In embodiments, a sample handling system may use a
pipette or similar device, with or without a pipette tip, to
aspirate, dispense, mix, transport, or otherwise handle the fluid.
The fluid may be dispensed from the sample handling system when
desired. The fluid may be contained within a pipette tip prior to
being dispensed, e.g., from an orifice in the pipette tip. In
embodiments, or instances during use, all of the fluid may be
dispensed; in other embodiments, or instances during use, a portion
of the fluid within a tip may be dispensed. A pipette may
selectively aspirate a fluid. The pipette may aspirate a selected
amount of fluid. The pipette may be capable of actuating stirring
mechanisms to mix the fluid within the tip or within a vessel. The
pipette may incorporate tips or vessels creating continuous flow
loops for mixing, including of materials or reagents that are in
non-liquid form. A pipette tip may also facilitate mixture by
metered delivery of multiple fluids simultaneously or in sequence,
such as in 2-part substrate reactions.
[0159] A sample handling system may include one or more fluidically
isolated or hydraulically independent units. For example, the
sample handling system may include one, two, or more pipette tips.
The pipette tips may be configured to accept and confine a fluid.
The tips may be fluidically isolated from or hydraulically
independent of one another. The fluid contained within each tip may
be fluidically isolated or hydraulically independent from one
fluids in other tips and from other fluids within the device. The
fluidically isolated or hydraulically independent units may be
movable relative to other portions of the device and/or one
another. The fluidically isolated or hydraulically independent
units may be individually movable. A sample handling system may
comprise one or more base or support. A base or support may support
one or more pipette or pipette units. A base or support may connect
one or more pipettes of the sample handling system to one
another.
[0160] A sample processing device may be configured to perform
processing steps or actions on a sample obtained from a subject.
Sample processing may include sample preparation, including, e.g.,
sample dilution, division of a sample into aliquots, extraction,
contact with a reagent, filtration, separation, centrifugation, or
other preparatory or processing action or step. A sample processing
device may be configured to perform one or more sample preparation
action or step on the sample. Optionally, a sample may be prepared
for a chemical reaction and/or physical processing step. A sample
preparation action or step may include one or more of the
following: centrifugation, separation, filtration, dilution,
enriching, purification, precipitation, incubation, pipetting,
transport, chromatography, cell lysis, cytometry, pulverization,
grinding, activation, ultrasonication, micro column processing,
processing with magnetic beads, processing with nanoparticles, or
other sample preparation action or steps. For example, sample
preparation may include one or more step to separate blood into
serum and/or particulate fractions, or to separate any other sample
into various components. Sample preparation may include one or more
step to dilute and/or concentrate a sample, such as a biological
sample, e.g., of blood, urine, sputum, material obtained from a
nasal swab, a throat swab, a cheek swab, or other sample, or other
biological samples. Sample preparation may include adding an
anti-coagulant or other ingredients to a sample. Sample preparation
may also include purification of a sample. In embodiments, all
sample processing, preparation, or assay actions or steps are
performed by a single device. In embodiments, all sample
processing, preparation, or assay actions or steps are performed
within a housing of a single device. In embodiments, most sample
processing, preparation, or assay actions or steps are performed by
a single device, and may be performed within a housing of a single
device. In embodiments, many sample processing, preparation, or
assay actions or steps are performed by a single device, and may be
performed within a housing of a single device. In embodiments,
sample processing, preparation, or assay actions or steps may be
performed by more than one device.
[0161] A sample processing device may be configured to run one or
more assay on a sample, and to obtain data from the sample. An
assay may include one or more physical or chemical treatments, and
may include running one or more chemical or physical reactions. A
sample processing device may be configured to perform one, two or
more assays on a small sample of bodily fluid. One or more chemical
reaction may take place on a sample having a volume, as described
elsewhere herein. For example one or more chemical reaction may
take place in a pill having less than femtoliter volumes. In an
instance, the sample collection unit is configured to receive a
volume of the bodily fluid sample equivalent to a single drop or
less of blood or interstitial fluid. In embodiments, the volume of
a sample may be a small volume, where a small volume may be a
volume that is less than about 1000 .mu.L, or less than about 500
.mu.L, or less than about 250 .mu.L, or less than about 150 .mu.L,
or less than about 100 .mu.L, or less than about 75 .mu.L, or less
than about 50 .mu.L, or less than about 40 .mu.L, or less than
about 20 .mu.L, or less than about 10 .mu.L, or other small volume.
In embodiments, all sample assay actions or steps are performed on
a single sample. In embodiments, all sample assay actions or steps
are performed by a single device. In embodiments, all sample assay
actions or steps are performed within a housing of a single device.
In embodiments, most sample assay actions or steps are performed by
a single device, and may be performed within a housing of a single
device. In embodiments, many sample assay actions or steps are
performed by a single device, and may be performed within a housing
of a single device. In embodiments, sample processing, preparation,
or assay actions or steps may be performed by more than one
device.
[0162] A sample processing device may be configured to perform a
plurality of assays on a sample. For example, a sample processing
device may be configured to detect, or to identify, or to measure
pathogen-identifying material in a sample. In embodiments, a sample
processing device may be configured to perform a plurality of
assays on a single sample. In embodiments, a sample processing
device may be configured to perform a plurality of assays on a
single biological sample, where the biological sample is a small
sample. For example, a small sample may have a sample volume that
is a small volume of less than about 1000 .mu.L, or less than about
500 .mu.L, or less than about 250 .mu.L, or less than about 150
.mu.L, or less than about 100 .mu.L, or less than about 75 .mu.L,
or less than about 50 .mu.L, or less than about 40 .mu.L, or less
than about 20 .mu.L, or less than about 10 .mu.L, or other small
volume. A sample processing device may be capable of performing
multiplexed assays on a single sample. A plurality of assays may be
run simultaneously; may be run sequentially; or some assays may be
run simultaneously while others are run sequentially. One or more
control assays and/or calibrators (e.g., including a configuration
with a control of a calibrator for the assay/tests) can also be
incorporated into the device; control assays and assay on
calibrators may be performed simultaneously with assays performed
on a sample, or may be performed before or after assays performed
on a sample, or any combination thereof. In embodiments, all sample
assay actions or steps are performed by a single device. In
embodiments, all of a plurality of assay actions or steps are
performed within a housing of a single device. In embodiments, most
sample assay actions or steps, of a plurality of assays, are
performed by a single device, and may be performed within a housing
of a single device. In embodiments, many sample assay actions or
steps, of a plurality of assays, are performed by a single device,
and may be performed within a housing of a single device. In
embodiments, sample processing, preparation, or assay actions or
steps may be performed by more than one device.
[0163] In embodiments, all of a plurality of assays may be
performed in a short time period. In embodiments, such a short time
period comprises less than about three hours, or less than about
two hours, or less than about one hour, or less than about 40
minutes, or less than about 30 minutes, or less than about 25
minutes, or less than about 20 minutes, or less than about 15
minutes, or less than about 10 minutes, or less than about 5
minutes, or less than about 4 minutes, or less than about 3
minutes, or less than about 2 minutes, or less than about 1 minute,
or other short time period.
[0164] A sample processing device may be configured to detect one
or more signals relating to the sample. A sample processing device
may be configured to identify one or more properties of the sample.
For instance, the sample processing device may be configured to
detect the presence or concentration of one analyte or a plurality
of analytes or a disease condition in the sample (e.g., in or
through a bodily fluid, secretion, tissue, or other sample).
Alternatively, the sample processing device may be configured to
detect a signal or signals that may be analyzed to detect the
presence or concentration of one or more analytes (which may be
indicative of a disease condition) or a disease condition in the
sample. The signals may be analyzed on board the device, or at
another location. Running a clinical test may or may not include
any analysis or comparison of data collected.
[0165] A chemical reaction or other processing step may be
performed, with or without the sample. Examples of steps, tests, or
assays that may be prepared or run by the device may include, but
are not limited to immunoassay, nucleic acid assay, receptor-based
assay, cytometric assay, colorimetric assay, enzymatic assay,
electrophoretic assay, electrochemical assay, spectroscopic assay,
chromatographic assay, microscopic assay, topographic assay,
calorimetric assay, turbidmetric assay, agglutination assay,
radioisotope assay, viscometric assay, coagulation assay, clotting
time assay, protein synthesis assay, histological assay, culture
assay, osmolarity assay, and/or other types of assays,
centrifugation, separation, filtration, dilution, enriching,
purification, precipitation, pulverization, incubation, pipetting,
transport, cell lysis, or other sample preparation action or steps,
or combinations thereof. Steps, tests, or assays that may be
prepared or run by the device may include imaging, including
microscopy, cytometry, and other techniques preparing or utilizing
images. Steps, tests, or assays that may be prepared or run by the
device may further include an assessment of histology, morphology,
kinematics, dynamics, and/or state of a sample, which may include
such assessment for cells.
[0166] A device may be capable of performing all on-board steps
(e.g., steps or actions performed by a single device) in a short
amount of time. A device may be capable of performing all on-board
steps on a single sample in a short amount of time. For example,
from sample collection from a subject to transmitting data and/or
to analysis may take about 3 hours or less, 2 hours or less, 1 hour
or less, 50 minutes or less, 45 minutes or less, 40 minutes or
less, 30 minutes or less, 20 minutes or less, 15 minutes or less,
10 minutes or less, 5 minutes or less, 4 minutes or less, 3 minutes
or less, 2 minutes or less, or 1 minute or less. The amount of time
from accepting a sample within the device to transmitting data
and/or to analysis from the device regarding such a sample may
depend on the type or number of steps, tests, or assays performed
on the sample. The amount of time from accepting a sample within
the device to transmitting data and/or to analysis from the device
regarding such a sample may take about 3 hours or less, 2 hours or
less, 1 hour or less, 50 minutes or less, 45 minutes or less, 40
minutes or less, 30 minutes or less, 20 minutes or less, 15 minutes
or less, 10 minutes or less, 5 minutes or less, 4 minutes or less,
3 minutes or less, 2 minutes or less, or 1 minute or less.
[0167] A device may be configured to prepare a sample for disposal,
or to dispose of a sample, such as a biological sample, following
processing or assaying of a sample.
[0168] In embodiments, a sample processing device may be configured
to transmit data obtained from a sample. In embodiments, a sample
processing device may be configured to communicate over a network.
A sample processing device may include a communication assembly
that may interface with the network. A sample processing device may
be connected to the network via a wired connection or wirelessly.
The network may be a local area network (LAN) or a wide area
network (WAN) such as the Internet. In embodiments, the network may
be a personal area network. In embodiments, the network may be a
virtual private network (VPN). The network may include the cloud.
The sample processing device may be connected to the network by
wire, cable, fiber optical, microwave, infrared, acoustic, or other
connections or connection means. The sample processing device may
be connected to the network without requiring an intermediary
device, or an intermediary device may be required to connect a
sample processing device to a network. A sample processing device
may communicate over a network with another device, which may be
any type of networked device, including but not limited to a
personal computer, server computer, or laptop computer; personal
digital assistants (PDAs) such as a Windows CE device; phones such
as cellular phones, smartphones (e.g., iPhone, Android, Blackberry,
etc.), or location-aware portable phones (such as GPS); a roaming
device, such as a network-connected roaming device; a wireless
device such as a wireless email device or other device capable of
communicating wireless with a computer network; or any other type
of network device that may communicate possibly over a network and
handle electronic transactions. Such communication may include
providing data to a cloud computing infrastructure or any other
type of data storage infrastructure which may be accessed by other
devices.
[0169] A sample processing device may provide data regarding a
sample to, e.g., a health care professional, a health care
professional location, such as a laboratory, or an affiliate
thereof. One or more of a laboratory, health care professional, or
subject may have a network device able to receive or access data
provided by the sample processing device. A sample processing
device may be configured to provide data regarding a sample to a
database. A sample processing device may be configured to provide
data regarding a sample to an electronic medical records system, to
a laboratory information system, to a laboratory automation system,
or other system or software. A sample processing device may provide
data in the form of a report.
[0170] A laboratory, device, or other entity or software may
perform analysis on data regarding a sample in real-time. A
software system may perform chemical analysis and/or pathological
analysis, or these could be distributed amongst combinations of
lab, clinical, and specialty or expert personnel. Analysis may
include qualitative and/or quantitative evaluation of a sample.
Data analysis may include a subsequent qualitative and/or
quantitative evaluation of a sample. Optionally, a report may be
generated based on raw data, pre-processed data, or analyzed data.
Such a report may be prepared so as to maintain confidentiality of
the data obtained from the sample, the identity and other
information regarding the subject from whom a sample was obtained,
analysis of the data, and other confidential information. The
report and/or the data may be transmitted to a health care
professional. Data obtained by a sample processing device, or
analysis of such data, or reports, may be provided to a database,
an electronic medical records system, to a laboratory information
system, to a laboratory automation system, or other system or
software.
[0171] Embodiments of the devices, systems, and methods disclosed
herein are discussed in the examples presented below.
EXAMPLES
[0172] The following examples disclose devices comprising a
sonicator effective to disrupt pathogens in a biological
sample.
Example 1 Systems for Disrupting Pathogens Comprising a
Sonicator
[0173] Embodiments of systems disclosed herein include devices
which have a sonicator configured to apply ultrasonic energy to a
sample solution. Application of ultrasonic energy to a sample
solution containing a pathogen may be effective to disrupt the
pathogen and to release pathogen-identifying material into the
solution, where it can be detected and the pathogen identified. In
embodiments, the amounts of pathogens present in the sample
solution may be quantified following application of ultrasonic
energy effective to disrupt the pathogen and to release
pathogen-identifying material into the solution. A schematic
illustration representing elements of an embodiment of a system
comprising devices having a sonicator is shown in FIG. 1
[0174] In embodiments, a tip portion of a sonicator of a device of
a system as disclosed herein be configured to improve the transfer
of ultrasonic energy to a sample solution, or to provide better
contact between the sonicator and a vessel containing a sample
solution, or both. In embodiments, a sonicator of a device of a
system as disclosed herein may have a sonicator horn; such a
sonicator horn may have a tip portion, which may be configured to
improve the transfer of ultrasonic energy to a sample solution, or
to provide better contact between a sonicator and a vessel
containing a sample solution, or both. In embodiments of systems
disclosed herein, a system may include a vessel configured to hold
a sample, and may include a vessel holder configured to hold a
vessel. A sample holder may be configured to place a vessel in a
proper position for sonication; may be configured to hold a vessel
during sonication (in embodiments, such holding may comprise
application of force effective to restrain a vessel during
sonication, e.g., to reduce loss of energy due to unwanted motion
of a vessel, or for improved energy transfer during sonication); or
may be configured to retain a vessel in place until further
operation is needed following sonication.
[0175] FIG. 1 provides an illustration of several components of an
exemplary system as disclosed herein. A system 10 may include a
controller 12; a power supply 16; a sonicator 18 (having a
sonicator horn 20, with a tip portion 21); a sample handling system
14; a vessel holder 26, configured to hold a vessel 22 (which may
have a cap 24); a detector 30; and a communication assembly 32. A
communication assembly 32 may be in communication with external
devices, or with a user, or with a laboratory, or with a network
(e.g., a computer network, such as a local area network, a wide
area network, the cloud, etc.), or other communication
destination.
[0176] As shown in FIG. 1, in embodiments, a device of a system 10
as disclosed herein may comprise a sample handling system 14
configured to place a sample within a vessel 22, by, for example,
delivering at least a portion of a sample to a vessel 22 and
depositing the at least a portion into a chamber of the vessel 22.
A sample handling system 14 may also be configured to place a
reagent, or other agent or material, within a vessel 22, by, for
example, delivering the reagent, other agent, or material, to a
vessel and depositing it into a chamber of the vessel.
[0177] As shown in FIG. 1, in embodiments, a device of a system 10
as disclosed herein may comprise a detector 30 for detecting
pathogen-identifying material from a sample. A detector 30 may be
any detector effective to detect the presence of
pathogen-identifying material in a sample. Such detection may be
aided by use of a label or other readily detectable agent, which
may be used in conjunction with a reagent that specifically binds
to, or reacts with, a pathogen-identifying material.
[0178] As shown in FIG. 1, in embodiments, a system 10 as disclosed
herein may comprise a controller 12 to operate the components of
the system. In embodiments, a device of a system 10 as disclosed
herein may comprise a controller 12 to operate components of the
device. A controller 12 may comprise a processor, or other
component, device, or element effective to oversee and control the
operation of a device. A controller may further comprise
communication components, devices, or elements effective to provide
communication with and between the controller 12 and other
components and elements of the device or system. Such internal
communication linkages are illustrated in FIG. 1 by dotted lines.
Note that, as shown, these internal communication linkages include
a quick release connection 28, allowing easy and rapid connection
and disconnection of these linkages; for example, such a linkage
(e.g., a quick release connection 28) may be useful in quickly
adding, or quickly removing, a sonicator 18 from a device, or for
quickly swapping one sonicator 18 for another sonicator 18 in a
device. In embodiments, such a quick release connection may be
useful in "hot swapping" of a sonicator 18 (connecting a sonicator,
removing a sonicator, or replacing one sonicator by another
sonicator during use of the sonicator or of other components, or
without requirement that the remainder of the device or system be
shut down for such changes). In embodiments, such linkages may be
direct connections lacking a quick release connection 28, or may
have other, less-readily disconnected, linkages. It will be
understood that, in embodiments of the present devices and systems,
other communication linkages may also be provided.
[0179] In embodiments, a system 10 as disclosed herein may comprise
a communication assembly 32, which may comprise one or more
communications link(s). In embodiments, a device of a system 10 as
disclosed herein may comprise a communication assembly 32, which
may comprise one or more communications link(s). Such a
communication assembly may comprise a display, for receiving
instructions and information, and for providing information and
data to a user, or to other components or equipment (illustrated by
the element numbered 34, which may represent one or more of, e.g.,
a user, another device, a laboratory, a network, the cloud, or
other communication destination). In embodiments, such a
communication assembly 32 may comprise a one-way communication link
(e.g., from the device or system to an element 34, which may
comprise a user, another device, a laboratory, a network, the
cloud, or other communication destination; or to the device or
system from an element 34 which may comprise a user, another
device, a laboratory, a network, the cloud, or other communication
source) and, in embodiments, may include a two-way (or
multiple-way) communication link(s), e.g., between the device or
system and a user, another device, a laboratory, a network, the
cloud, or other communication targets, as represented by element
34. Such communication links with communication destinations or
sources are indicated by the dotted arrows in the figure.
[0180] In embodiments, a device or system 10 as disclosed herein
may comprise a power supply 16 to enable the operation of the
components. Power connections are illustrated in FIG. 1 by solid
lines. It will be understood that, in embodiments of the present
devices and systems, other power connections may also be provided.
Note also that, as shown, the power connection to the sonicator 18
includes a quick release connection 28, allowing easy and rapid
connection and disconnection of that connection. Thus, for example,
such a quick release connection 28 may be useful in quickly adding,
or quickly removing, a sonicator 18 from a device, or for quickly
replacing a sonicator 18 in a device. In embodiments, such a quick
release connection 28 may be useful in "hot swapping" of a
sonicator 18 (connecting a sonicator, removing a sonicator, or
replacing one sonicator by another sonicator during use, or without
requirement that the remainder of the device or system be shut down
for such changes). In embodiments, such a power connection may be a
direct connection lacking a quick release, or may have other,
less-readily disconnected, linkages. It will also be understood
that, in embodiments, a power connection to a sonicator 18 may
serve as the control connection (or one of the control connections)
that controls the frequency, cycling, or other aspects of sonicator
operation. A control connection may serve to turn-off the
high-voltage power supply 16 to a sonicator 18 during (and
following) disconnection of the sonicator 18 to prevent delivery of
a shock to personnel or to prevent shorting power to an improper
location. In embodiments, a control connection may include a quick
release connection 28.
[0181] As disclosed herein, ultrasonic energy may be applied to a
sample by direct contact between the sample and a sonicator 18
(e.g., by immersion of a sonicator tip or tip portion 21 in a
sample solution). As disclosed herein, ultrasonic energy may be
applied indirectly to a sample, by contacting a sonicator tip
portion or tip 21 with a material separating the sonicator tip 21
and the sample solution, where the material is in contact both with
the sonicator tip portion or tip 21, and with the sample solution.
In embodiments comprising indirect contact, the material separating
the sonicator 18 and the solution may comprise a compliant barrier.
In embodiments comprising indirect contact, the material separating
the sonicator 18 and the solution may comprise a wall of a vessel
containing a sample. In any embodiment comprising indirect contact,
the material separating the sonicator 18 and the solution is
configured to transmit ultrasonic energy from the sonicator tip
portion to the sample solution.
Example 2 Sonicators and Vessels
[0182] Examples of sonicators suitable for use in the present
devices, systems and methods are shown in FIG. 2 and FIG. 3. FIG. 2
may provide an illustration of an embodiment in which a distal
portion of the sonicator 40 (e.g., the tip 44 of the sonicator horn
42, shown within vessel 46), termed in this example the "tip" 44,
contacts a sample solution in a vessel 46. In such embodiments, the
tip 44 contacts the sample solution. Direct contact with the sample
solution provides for direct transfer of energy from the sonicator
40 to the sample solution. However, such direct contact requires
that the tip 44 touch or enter into the sample solution, providing
a possible source of contamination of the sample (and subsequent
samples) and requiring cleaning steps and/or coating of the tip 44
to reduce such contamination.
[0183] FIG. 2 may also provide an illustration of an embodiment in
which the tip 44 is separated from the sample solution by a
compliant barrier (such a compliant barrier conforms, to a greater
or lesser degree, to the outline of the tip 44 as the tip 44 is
pressed into a vessel 46 towards a solution within the vessel 46).
In such an embodiment, the tip 44 contacts the barrier, and the
barrier contacts a sample solution in a vessel 46. In such
embodiments, the tip 44 does not contact the sample solution,
avoiding possible problems which may arise due to direct contact
with the sample solution. Barrier materials tested include latex,
nitrile, and polyurethane barriers. However, a compliant barrier
may break or become porous during use, and may absorb ultrasonic
energy, reducing the efficiency of transferring ultrasonic energy
from a tip 44 to a solution.
[0184] An alternative embodiment is shown in FIG. 3, showing a
sonicator 50 having a sonicator horn 52 and sonicator tip 54 in
contact with a vessel 56 having a flat face 58 on a wall of the
vessel 56. FIG. 3 shows an example in which a distal portion of the
sonicator 50 contacts a flat face 58 of the vessel 56 holding a
sample solution; flat face 58 is configured to be complementary to
sonicator tip 54 for efficient energy transfer between the
sonicator and the vessel and its contents. In such embodiments, no
portion of the sonicator 50 contacts the sample solution,
preventing contamination of the sample and of the sonicator 50, so
that there is no requirement to clean or condition the sonicator
tip 54 before its re-use. A wall 58 of a vessel, which may be more
rigid, and less compliant than a compliant barrier, may be more
efficient at transferring ultrasonic energy from a tip 54 to a
solution than a compliant barrier; for example, a rigid wall may
absorb less energy than would be absorbed by a compliant
barrier.
[0185] Although the contact between sonicator 50 and sample
solution is indirect in embodiments comprising the configuration
illustrated in FIG. 3, ample ultrasonic energy is transferred via
the wall of a vessel 56 into a sample solution contained therein,
and cells, including pathogen cells, may be, and have been,
disrupted in this way. Ultrasonic energy transfer from sonicator 50
to sample solution is improved in this configuration by providing a
flat surface 58 on a wall of the vessel 56, as disclosed herein,
which is complementary to a flat surface on the sonicator tip 54.
Transmission of ultrasonic energy in the configuration illustrated
in FIG. 3 is improved by tight contact between a sonicator tip 54
and a wall of a vessel 56. Application of force between a wall of a
vessel 56 (particularly a flat face 58 of a vessel 56) and the
sonicator tip 54 is also helpful to provide effective ultrasonic
energy transfer from sonicator 50 to sample solution.
[0186] Forces of between 1 newton (N) and 16 N have been examined.
For example, experiments in which forces of 1, 2, 3, 4, 6, and 10 N
were applied to force contact between the tip of a sonicator horn
and a polystyrene tube resulted in effective disruption of a test
pathogen. In these experiments, 332 .mu.L of solution containing E.
coli (ATCC 884) bacteria was held in a polystyrene tube. A
sonicator probe was applied to the outer wall of the polystyrene
tube with 1, 2, 3, 4, 6, and 10 N of force and ultrasonic energy
was applied for 10 seconds. Such disruption was comparable to that
obtained with a commercial sonicator in which vials are placed in
wells in a block connected to an ultrasonic generator. However, the
results with 1 N of contact force were variable, and considered
inferior to the results obtained with the commercial sonicator.
From the results of these and other experiments, it is believed
that a contact force of between about 2 N to about 10 N, or between
about 2 N to 6 N, or between about 3 N to about 5 N, or about 4 N,
is useful for providing effective transfer of ultrasonic energy for
disruption of pathogens by contacting a wall of a vessel containing
a sample solution.
[0187] Different materials exhibit different ultrasonic energy
transfer characteristics. For example, preferred vessel materials
for ultrasonic energy transfer through a vessel wall include
polystyrene, polycarbonate, and polyethylene. Polystyrene was found
to be a better material than polycarbonate and polyethylene;
polycarbonate was found to be a better material than
polyethylene.
[0188] Ultrasonic disruption of pathogens may be performed with
varying frequencies of ultrasonic energy. Higher frequencies allow
for shorter sonicator horns; conversely, sonicators designed to
provide ultrasonic energy at lower frequencies will have longer
sonicator horns than do sonicators designed for higher frequencies.
For example, ultrasonic frequencies suitable for use in devices,
systems and methods disclosed herein include ultrasonic frequencies
of from about 20 kHz to about 60 kHz, or from about 20 kHz to about
50 kHz, or from about 20 kHz to about 40 kHz. In embodiments of the
devices, systems and methods disclosed herein include ultrasonic
frequencies devices, systems and methods disclosed herein, suitable
ultrasonic frequencies include frequencies of about 20 kHz, about
25 kHz, about 28 kHz, about 30 kHz, about 35 kHz, about 40 kHz,
about 45 kHz, about 50 kHz, about 55 kHz, and about 60 kHz.
[0189] A sonicator may include or have attached to it multiple
elements. FIG. 4 shows an example of an embodiment of a sonicator
60 having a transducer 62, a motor 64, a drawbar 66, a horn 68, and
a collet 70. In embodiments, a motor 64 may be effective to move
(e.g., rotate) a drawbar 66 connected to a collet 70 at the tip of
the sonicator 60. As shown in FIG. 4, such a sonicator 60 may have
a hollow sonicator horn 68; in the embodiment shown, the hollow
sonicator horn 68 encloses, and allows passage of, a drawbar 66
which provides functional connection between a collet 70 at the
distal end of the sonicator 60 and transducer 62 (and thus with the
sonicator body as well). A drawbar 66 such as the drawbar 66 shown
in the figure that is connected to a collet 70 and a motor 64 may
be used to cinch a collet 70 around a vessel contained within the
collet 70 to provide a tight fit between the collet 70 and vessel;
such a tight fit may aid in the efficient transfer of ultrasonic
energy between the transducer 62 of the sonicator 60 (via the
collet 70) and the vessel. As shown in the figure, a sonicator 60
having such a horn 68 and drawbar 66 may be configured to transfer
ultrasonic energy from its proximal end (shown here adjacent the
motor 64, near the transducer 62) to its distal end (adjacent the
collet 70). A collet 70 such as the collet 70 shown in the figure
may be configured to mate with and hold a vessel effective to
provide ultrasonic energy to a sample within a vessel. In
embodiments, a sonicator 60 having such components at its tip may
be configured to transfer ultrasonic energy to a sample solution,
and in embodiments, may be configured to transfer ultrasonic energy
to a sample solution via a vessel wall.
[0190] As shown in FIG. 5 depicting an exemplary sonicator assembly
100, a sonicator 102 may be mounted on a movable mount 106 for use
in an embodiment of the devices disclosed herein. The sonicator
mount 106 shown in FIG. 5 is operably connected to a solenoid 120,
effective that when the solenoid 120 is activated, the sonicator
mount 106, and its attached sonicator 102, are drawn towards the
solenoid 120 (in a distal direction with reference to the sonicator
102). This motion moves the sonicator horn 104 in the same (distal)
direction, moving the tip of the sonicator horn 104 deeper into the
sonicator horn port 114 of the vessel holder 112. When a vessel is
in place within the vessel holder 112, activation of the solenoid
120 and the resulting movement presses the tip of the sonicator
horn 104 into contact with a side wall of the vessel. In
embodiments, the action of the solenoid 120 generates a force at
the area of contact between the tip of the sonicator horn 104 and
the vessel wall. Such a force may be, for example, between about 2
N and about 10 N, or between about 2 N and about 6 N, or preferably
between about 3 N and about 5 N, or, in embodiments, about 4 N of
force.
[0191] As shown, a vessel holder 112 may have a further aperture
116 in a vessel holder wall 118, allowing access to another portion
of the vessel, and including optical access without possible
interference by the vessel holder 112. As shown, a vessel holder
112 may have walls 118 without apertures; such walls 118 may be
configured for optical access as well, by construction using
suitable materials; by providing a wall of proper flatness,
orientation, or thickness of the wall; by proper preparation or
construction of the wall surface, or by other means. As shown in
FIG. 5, in embodiments, a power connection 110 to a sonicator 102
may also serve as a control connection (or one of the control
connections) that controls the frequency, cycling, or other aspects
of sonicator operation. Note also that a control/power connection
110 to the sonicator 102 may include a quick release connection 108
(as illustrated in FIG. 5), allowing easy and rapid connection and
disconnection of that connection; for example, such a quick release
connection 108 may be useful in adding, removing, or replacing a
sonicator 102. In embodiments, such a quick release connection may
be useful in "hot swapping" of a sonicator 102. In embodiments,
such a control/power connection 110 may be a direct connection
lacking a quick release, or may have other, less-readily
disconnected, linkages.
[0192] FIGS. 6A-6B provide further illustration of a sonicator
assembly 100, and the design and operation of a movable sonicator
102 operably connected to a solenoid 120 for moving a sonicator
horn 104 and its tip into functional contact with a wall of a
vessel containing a sample solution. FIGS. 6A and 6B present a side
view of an embodiment of a sonicator 102 mounted and operably
connected with a solenoid 120 that is configured to move the
sonicator horn 104 in a transverse direction. In FIG. 6A, the
solenoid is not activated. FIG. 6A shows a side view of an
embodiment of a sonicator 102 mounted with a solenoid 120
configured to move the sonicator horn 104 in a transverse direction
(e.g., leftwardly or rightwardly as shown in the figure). The oval
encloses an area of interest, in which the tip of the sonicator
horn 104 moves to and contacts a vessel wall when a vessel is in
place within the vessel holder. As shown, a spring 126 is provided
to urge the sonicator 102 away from the vessel holder 112 upon
release of solenoid force drawing the sonicator 102 towards the
vessel holder 112. The sonicator horn 104 is disposed so as to
approach a vessel held within the vessel holder 112 when the
sonicator 102 is moved to the right in the figure. At rest (with
the solenoid 126 off), as shown in FIG. 6A, the sonicator horn 104
does not contact a vessel held within the vessel holder 112, and
the spring 126 is in an extended conformation.
[0193] The solenoid 120 is activated in FIG. 6B, the sonicator horn
104 is in contact with an outer wall of a vessel held within the
vessel holder 112, and the spring 126 is in a compressed
conformation. Operation of the sonicator 102 in this configuration,
in which the sonicator horn 104 is in contact with a wall of a
vessel held within the vessel holder 112, is effective to provide
ultrasonic energy to a sample solution within the vessel. Transfer
of ultrasonic energy is improved by providing a firm contact
between the tip of the sonicator horn 104 and the vessel wall; as
discussed above, ultrasonic energy transfer is improved by
providing transverse force (towards the vessel wall) of between
about 2 N and about 10 N, or between about 2 N and about 6 N, or
about 3N to about 5N, or about 4 N of force. Providing sufficient
ultrasonic energy for a sufficient amount of time is effective to
disrupt cells within the sample solution; in particular, pathogen
cells within the sample solution may be disrupted, effective to
release pathogen-identifying material for detection,
identification, and measurement.
[0194] FIG. 7 provides a further, more detailed illustration of
contact between the tip of sonicator horn 104 and a vessel wall.
Also shown are sonicator mount 106, spring 126, base 125 with
attachment mechanism 122, and a clamp 128 for securing a sonicator
102 to a sonicator mount 106. The vessel 130 is held in the vessel
holder 112; in this example, the vessel 130 has a vessel cap 132.
In embodiments, a vessel 130 may be filled with a sample solution
prior to placement of the vessel cap 132. In embodiments, a vessel
130 may be filled with sample solution through a vessel cap 132,
e.g., by a conduit through the vessel cap 132. Such a conduit may
be temporary (e.g., may be provided by a hollow needle piercing the
cap 132, where the path of the needle may reseal following removal
of the needle from the cap 132). Such a conduit may be permanent,
e.g., may be a channel or tube that is a permanent feature of the
vessel cap 132. In embodiments, such a conduit may itself be
capped. In embodiments, the vessel 130 may be filled by the sample
handling system. In embodiments, the cap 132 may be placed on the
vessel 130 by the sample handling system following filling of the
vessel 130. In embodiments, the cap 132 may be removed by the
sample handling system; for example, in embodiments, the cap 132
may be removed by the sample handling system prior to filling the
vessel 130 with sample solution.
[0195] A cap 132 as shown in FIG. 7 may be effective to prevent
loss of sample fluid during sonication, and to prevent spread of
sample contents (by spillage, aerosolization, or other means)
outside the vessel. In this figure, a vessel 130 is shown held in a
vessel holder 112, and a sonicator horn 104 is shown in contact
with a side wall of the vessel 130. Such a configuration is an
operative configuration effective to disrupt cells within the
sample solution, such as pathogen cells within the sample
solution.
[0196] A schematic illustration of a device 200 (enclosed by a
device housing 202) having a sonicator 204 and other features as
disclosed herein is provided in FIG. 8. The device illustrated by
the figure has a sonicator 204 with a sonicator horn 208, which can
be made to contact a wall of a vessel 212 (e.g., by motion of a
movable sonicator mount 206, as indicated in FIGS. 6A, 6B, and 7).
The vessel 212 can contain a sample solution, and is configured to
be held by a vessel holder 210. The vessel 212 shown in FIG. 8 has
a vessel cap 214. A sample handling system 216 may be provided
effective to transport a biological sample, or a portion thereof,
and reagents if applicable, to the vessel 212. As discussed herein,
a sample handling system 216 may also be configured to manipulate
or move a vessel 212, a vessel cap 214, or other article, within a
housing of a device 200. A device 200 may have a detector 218,
e.g., within the device housing 202 as illustrated in the figure,
such as a detector 218 configured to detect pathogen-identifying
materials released by sonication of a sample solution. A device 200
may further include other components 220 for performing assays,
detecting analytes, and collecting and communicating data obtained
from biological samples, and other actions. Such other components
220 may include, e.g., components configured to perform assays, or
to mix reagents, cameras, light sources, lenses, filters,
temperature control devices; temperature, optical, chemical or
electronic sensors; mechanical components associated with such
components, within the device housing as illustrated in the
figure.
[0197] A sonicator 204 may be used to deliver ultrasonic energy to
a sample. In embodiments, a sonicator 204 may be used to deliver
ultrasonic energy to a sample in a vessel 212 by contacting a
vessel wall, and applying ultrasonic energy to the wall effective
to transfer ultrasonic energy to the sample within the vessel 212.
Contact between a sonicator 204 and a vessel wall is typically
between the sonicator horn 208, e.g., the tip of the sonicator horn
208, and a vessel wall; a portion of a vessel wall may be
configured or adapted for efficient contact with a tip of a
sonicator horn 208. The amount of energy provided by a sonicator
204 may depend on the ultrasonic frequency of the ultrasonic energy
applied. The amount of energy provided by a sonicator 204 may
depend on the amplitude of the displacement produced by the
ultrasonic energy. The amount of energy provided by a sonicator 204
may depend on the duration of the application of the ultrasonic
energy. The amount of energy provided may depend on the contact
force applied by the sonicator 204 to the vessel 212. The amount of
energy provided by a sonicator 204 may depend on the area of
contact between a sonicator 204 and a vessel 212. The amount of
energy provided by a sonicator 204 may depend on the duty cycle of
the application of ultrasonic energy (e.g., where the ultrasonic
energy is not applied continuously for a duration, but is applied
for a first time period, application of ultrasonic energy is ceased
for rest time period, and then ultrasonic energy is applied for a
second time period, the combination of such periods may be termed a
"duty cycle"; a duty cycle may be repeated to provide multiple
periods during which ultrasonic energy is applied, separated by
rest periods). The amount of energy provided by a sonicator 204 may
depend on the shape of the time-varying ultrasonic energy applied
(e.g., ultrasonic energy may be produced by a transducer driven by
square wave, sawtooth wave, sine wave, or other shaped signals).
The amount of energy applied by a sonicator 204 to a fluid within a
vessel contacted by the sonicator 204 may depend on the material,
or design, or thickness of a wall or walls of the vessel.
[0198] In embodiments, a sonicator may be used to disrupt a cell,
such as a pathogen cell in a sample. In embodiments, a sonicator
may be used to mix a solution; for emulsification of a solution or
a mixture of solutions; for resuspension of a material, e.g.,
resuspension of a sample following centrifugation of the sample;
for aerosolization of a liquid; to heat a solution; to disperse a
sample in, or into, a solution; to disintegrate a material; to
de-gas a liquid; and for other uses. The power requirements for one
use of a sonicator may differ from the power requirements of
another use of a sonicator. For example, the power required to
disrupt a cell in a solution may be greater than the power required
to mix a solution. The duration of power applied to a solution may
differ depending on the use of the sonicator; for example, the
duration of application of ultrasonic power used to heat a solution
will depend on the initial temperature of the solution, and the
desired final temperature. The shape of the control signal sent to
an ultrasonic transducer affects the power applied to a solution,
and some shapes may be more effective than others depending on the
use of the sonicator.
[0199] A device as disclosed herein may comprise more than one
sonicator. A system as disclosed herein may comprise more than one
sonicator. In an embodiment as shown in FIG. 9, a single power
supply may provide power to drive multiple sonicators, where the
power supply is connected via a multiplexer to the sonicators. In
the example shown in FIG. 9, a single 24 volt, 2 Amp (DC) power
supply 302 can output about 200 V to about 400 V (AC, at 40 Khz;
e.g., about 50 VRMS to about 300 VRMS) via a single 6-channel probe
power multiplexer 304 effective to distribute six multiplexed
output lines to six sonicators. In embodiments, such a
configuration may provide six devices, in a system comprising
multiple devices (e.g., where each device 308 and 310 as indicated
in FIG. 9 may be termed a "bay"). In embodiments, the output lines
may be detachable lines, indicated by the detachable plug 306,
configured to disengage a bay when desired or needed, and
configured to allow reconnection to the power supply 302 when
desired or needed (such connections 306 as indicated in the figure
may be termed "hot plugs"). A multiplexer 304 may be configured to
control the sonicator pulse duration, or amplitude, or both, and so
to control the duration of ultrasonic power transfer to a sample
solution. The operation of the multiplexer 304 selection may be
according to a set pattern or frequency, or may be controlled, for
example, by a controller as shown in FIG. 1, or may be controlled
by other means. In embodiments of devices and systems disclosed
herein, 2, 3, 4, 5, 6, 7, 8, 9, 10, or more sonicators may be
configured in this way, and may be controlled in this way.
[0200] In embodiments, a single sonicator may contact a plurality
of vessels. In embodiments in which a single sonicator contacts a
plurality of vessels, the vessels may be disposed in an array of
vessels. An array of vessels may be a linear array, in which a
sonicator may move along the array and contact vessels
sequentially. An array of vessels may be circular, or semicircular
array, in which a sonicator rotates around an axis effective to
position the sonicator tip for contact with vessels disposed in
such a circular or semicircular array. An array of vessels may
comprise a combination of linear and curved arrays, may have an
irregular shape, or vessesl may be disposed in any shaped array
suitable for contact with a sonicator tip.
[0201] In embodiments, multiple sonicators may contact a plurality
of vessels, e.g., with a first sonicator contacting a first vessel,
a second sonicator contacting a second vessel, and so forth. In
embodiments in which multiple sonicators contact a plurality of
vessels, the vessels may be disposed in an array of vessels. In
embodiments in which multiple sonicators contact a plurality of
vessels, the sonicators may be disposed in an array of sonicators.
An array of sonicators, an array of vessels, and both arrays, may
be a linear array, may be a curved array, may be a combination of
such arrays, may be an irregular array, and may be any shaped array
suitable for contact between a sonicator tip and a vessel.
[0202] An embodiment of a vessel suitable for containing a sample
solution for sonication is shown in several views in FIG. 10. A
side view of a vessel 502 having mating sockets 504, with a flat
wall surface 510 facing outward, a flat bottom 506, and an opening
508 (for filling the vessel) shown at the top is provided in FIG.
10A. A side view with a flat wall surface 510 facing rightward is
shown in FIG. 10B while FIG. 10C shows an angled view of the vessel
502. FIG. 10D shows a cross-sectional view of a vessel 502. The
vessel 502 of this embodiment has a flat side wall 510 and a flat
bottom 506; both the flat side wall 510 and the flat bottom 506 are
configured to make effective contact with a tip portion of a
sonicator, such as a tip of a sonicator horn to provide for
transfer of ultrasonic energy from the sonicator to a sample
solution within the vessel. The wider portion at the top of the
vessel includes surfaces and mating sockets 504. The mating sockets
504 comprise recesses configured for engagement of a transport
and/or force-providing member (e.g., a nozzle of a sample handling
system) which a) allows transport of the vessel 502 and b) provides
a surface for provision of downward force to oppose upward force of
a sonicator horn placed on a flat bottom surface 506 of the vessel.
Openings of the mating sockets 504 are visible in this view, as is
an inner ridge 514 within the internal chamber 512 of the vessel.
In embodiments, mating sockets 504 may be configured to mate with a
mechanical component configured to transport a vessel, or to apply
force to a vessel, or both. In embodiments, such a mechanical
component may be, or may comprise, a sample handling system. In
embodiments, such a mechanical component may comprise a pipette, a
nozzle, or other mechanical component.
[0203] FIG. 11 provides further embodiments of vessels suitable for
containing a sample solution for sonication. Embodiments of vessels
602 with caps 604 shown in FIG. 11 have rounded bottoms 606, which
are believed to be more suitable for mixing, re-suspension, and
other applications in which a sonicator is applied to a wall of a
vessel. FIGS. 11A, 11C, and 11D show embodiments of tubular vessels
with rounded bottoms 606. FIG. 11B shows an embodiment of a conical
vessel 602 with a rounded bottom 606. FIG. 11C an embodiment of an
elongated tubular vessel 602 with a rounded bottom 606. FIG. 11D
shows an embodiment of a wide tubular vessel 602 with a rounded
bottom 606. The vessels 602 shown in FIG. 11 are shown with caps
604; it will be understood that a vessel 602 may not be provided
with a cap 604. In embodiments, caps 604 may be removable. In
embodiments, caps 604 may not be removable, but may be piercable
effective to fill a vessel 602 without removing the cap 604. In
embodiments, caps 604 may not be removable, but may comprise a
passage for filling or dispensing.
[0204] FIG. 12 provides yet further embodiments of vessels suitable
for containing a sample solution for sonication. FIG. 12A shows an
embodiment of a tubular vessel 602 with a rounded bottom 606 and a
tapered (e.g., partially conical-shaped cap) cap 604. A cap 604 as
shown in FIG. 12 A may form a tight seal with the lip 610 of the
vessel 602 when pressed downwardly, due to its partially conical
shape. A cap 604 as shown in FIG. 12 A may protrude into the
interior of the vessel 602, or in embodiments, may protrude into
the vessel 602 only a small amount, or in embodiments, may not
protrude into the vessel 602. FIG. 12B shows an embodiment of a
conical vessel 602 with a rounded bottom 606 having flat side
surfaces 608 configured to engage with a sonicator tip. As
indicated by the example of the embodiments of FIG. 11B and FIG. 12
B, a conical vessel 602 may have one, two, three or more flat
surfaces 608 configured for contact with a sonicator. In
embodiments, a conical vessel 602 may have no such flat surfaces
608, but a sonicator tip may press against the outer wall of a
conical vessel lacking such surfaces as indicated in FIG. 11B and
FIG. 12B. FIG. 12C shows an embodiment of an elongated tubular
vessel 602 with a rounded bottom 606 and a protruding flat surface
608 configured to engage with a sonicator tip. It will be
understood that, in embodiments, an elongated tubular vessel 602
with a rounded bottom 606 may have two, three, or more protruding
flat surfaces 608 configured to engage with a sonicator tip. FIG.
12D shows an embodiment of a wide tubular vessel 602 with a rounded
bottom 606 having a cap 604 connected to the vessel via cap linkage
612. Such a cap 604 may be removable, yet remain linked to the
remainder of the vessel 602. A cap linkage 612 may comprise a
hinge, a tab, a thread, a perforated tab, or any element effective
to connect the cap 604 with the body of the vessel 602 and allow
the opening and closing of the vessel 602 by movement of the cap
604.
[0205] As shown in FIG. 13, a vessel 816 having a flat bottom 824
may be contacted as illustrated by a tip 822 of a sonicator 802.
Such contact is effective to transfer ultrasonic energy to the wall
(i.e., flat bottom 824) of the vessel 816 and thereby to a sample
solution contained within the vessel 816, effective to disrupt
cells in the sample solution. In embodiments, a sonicator tip 822
may contact a wall of a vessel 816, such as a lower wall 824 as
illustrated in FIG. 13, while the vessel 816 is held in a vessel
holder. In embodiments, a vessel 816 held in a vessel holder may be
in contact with no other components that provide restraint or
retaining forces. For example, in embodiments, a sonicator tip 822
may contact a wall of a vessel 816, such as a lower wall 824 as
illustrated in FIG. 13, while the vessel 816 is held in a vessel
holder without contact of a nozzle with a mating socket 818.
[0206] In embodiments, functional contact between the vessel 816
and the sonicator 802 may be aided or improved by application of
force (as illustrated by the downward arrow 820 in the figure)
urging the vessel 816 onto the tip 822 of the sonicator horn 806,
improving functional contact and improving energy transfer from the
sonicator 802 to the sample solution and cells within it. Force
urging contact between the sonicator tip 822 and the vessel bottom
824 may be provided by a spring 812, e.g., a spring 812 as shown in
the figure. A spring 812 may be within, and in contact with, a
sonicator housing 804, as well as in contact with a sonicator 802;
a sonicator housing may have a bracket 814 for securing a sonicator
in place to or within a device. In embodiments, a nozzle of a
sample handling system, or two nozzles of a sample handling system,
may engage with one or both mating sockets 818, and apply downward
force effective to urge the base 824 of the vessel 816 onto the
sonicator tip 822 to improve contact between the vessel 816 and the
sonicator tip 822. In further embodiments, where one or two nozzles
of a sample handling system engage with one or both mating sockets
818, and apply downward force effective to urge the base 824 of the
vessel 816 onto the sonicator tip 822, such application of force
820 in conjunction with the presence of the spring 812 shown in
FIG. 13 may improve the contact, or improve control of the amount
or effect of the applied force. The action and function of the
spring 812 shown in FIG. 13 is to urge the sonicator 802 and vessel
816 together. This action is opposite to that of the springs 812
shown in FIGS. 6A, 6B, and 7, which act to urge the sonicator and
vessel apart from each other.
[0207] Thus, in embodiments, force 820 urging contact between a
sonicator tip 822 and a wall of a vessel 816 (e.g., a bottom wall
824) may be provided by one or more of a solenoid, a spring, a
sample handling system, or other means.
[0208] As shown in FIG. 13, in embodiments, a power/control
connection 808 to a sonicator 802 may serve both as a power
connection and as a control connection (or one of the control
connections) that control the frequency, cycling, or other aspects
of sonicator 802 operation. As shown in FIG. 13, a control/power
connection 808 to the sonicator 802 may include a quick release
connection 810. In other embodiments, such a control/power
connection 808 may be a direct connection lacking a quick release,
or may have other, less-readily disconnected, linkages.
[0209] Sonication is effective to disrupt cell membranes.
Sonication may be used to release pathogen-identifying material
into a sample solution effective to assay the sample, to detect the
presence of pathogens, to identify pathogens present in the
solution, and to measure the amount of pathogens present in the
solution. Pathogens tested using devices with sonicators as
disclosed herein included E. coli 884, S. pneumonia 3508, and
Adenovirus 1.
[0210] Since nucleic acids are enclosed by cell or viral membranes,
detection of target pathogens demonstrates that the sonication has
disrupted these membranes and made the pathogen-identifying nucleic
acids available for assay. As shown in FIG. 14, pathogens were
detected following sonication using quantitative polymerase chain
reaction (qPCR). qPCR assays detect target nucleic acids indicative
of the target pathogen; repeated cycles double the number of copies
of the target (if present); this process is termed amplification.
Other detection methods may also be used, including other PCR
methods known in the art, other nucleic acid assays known in the
art, ELISA assays, other antibody-based assays, hemagglutinin
assays (e.g., for influenza), and other assays. The horizontal axis
of FIG. 14 represents the number of cycles while the vertical axis
(in relative light units) represents the numbers of copies of the
target nucleic acid sequence indicative of the target pathogen. The
horizontal line near the bottom of the figure (labeled "Detection
Limit") indicates the lower limit for detection of pathogen nucleic
acids. The cycle number at which a trace passes this threshold is
the "cycle threshold" or "Ct" number; a target is considered
detected when the results pass the threshold. Control traces (the
two traces at the far-right of the figure, labeled "Control") are
shown, in which pathogen-containing solutions in polypropylene
containers mounted in wells of a block connected to an ultrasonic
generator were subjected to sonication). The other traces (a trace
labeled "a" and pairs of traces labeled "b" and "c") result from
experiments in which a sonicator tip contacted walls of vessels
holding pathogen-containing solutions. As shown in the figure,
nucleic acids from target pathogens were detectable after about 25
cycles. Thus, sonication as disclosed herein may be effective to
disrupt pathogens for use in the detection, identification, and
measurement of pathogens in a sample.
[0211] While the above is a complete description of the preferred
embodiment as described herein, it is possible to use various
alternatives, modifications and equivalents. Therefore, the scope
of the present invention should be determined not with reference to
the above description but should, instead, be determined with
reference to the appended claims, along with their full scope of
equivalents. Any feature, whether preferred or not, may be combined
with any other feature, whether preferred or not. The appended
claims are not to be interpreted as including means-plus-function
limitations, unless such a limitation is explicitly recited in a
given claim using the phrase "means for." It should be understood
that as used in the description herein and throughout the claims
that follow, the meaning of "a," "an," and "the" includes plural
reference unless the context clearly dictates otherwise. Also, as
used in the description herein and throughout the claims that
follow, the meaning of "in" includes "in" and "on" unless the
context clearly dictates otherwise. Finally, as used in the
description herein and throughout the claims that follow, the
meanings of "and" and "or" include both the conjunctive and
disjunctive and may be used interchangeably unless the context
expressly dictates otherwise. Thus, in contexts where the terms
"and" or "or" are used, usage of such conjunctions do not exclude
an "and/or" meaning unless the context expressly dictates
otherwise.
[0212] This document contains material subject to copyright
protection. The copyright owner (Applicant herein) has no objection
to the facsimile reproduction by anyone of the patent document or
the patent disclosure, as they appear in the US Patent and
Trademark Office patent file or records, but otherwise reserves all
copyright rights whatsoever. The following notice shall apply:
Copyright 2014 Theranos, Inc.
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