U.S. patent application number 15/996949 was filed with the patent office on 2018-10-04 for system and method for processing and detecting nucleic acids.
The applicant listed for this patent is NeuMoDx Molecular, Inc.. Invention is credited to Sundaresh Brahmasandra, Michael T. Kusner, Jeffrey Williams.
Application Number | 20180280978 15/996949 |
Document ID | / |
Family ID | 53264221 |
Filed Date | 2018-10-04 |
United States Patent
Application |
20180280978 |
Kind Code |
A1 |
Williams; Jeffrey ; et
al. |
October 4, 2018 |
SYSTEM AND METHOD FOR PROCESSING AND DETECTING NUCLEIC ACIDS
Abstract
A system and method for processing and detecting nucleic acids
from a set of biological samples, comprising: a capture plate and a
capture plate module configured to facilitate binding of nucleic
acids within the set of biological samples to magnetic beads; a
molecular diagnostic module configured to receive nucleic acids
bound to magnetic beads, isolate nucleic acids, and analyze nucleic
acids, comprising a cartridge receiving module, a heating/cooling
subsystem and a magnet configured to facilitate isolation of
nucleic acids, a valve actuation subsystem configured to control
fluid flow through a microfluidic cartridge for processing nucleic
acids, and an optical subsystem for analysis of nucleic acids; a
fluid handling system configured to deliver samples and reagents to
components of the system to facilitate molecular diagnostic
protocols; and an assay strip configured to combine nucleic acid
samples with molecular diagnostic reagents for analysis of nucleic
acids.
Inventors: |
Williams; Jeffrey; (Ann
Arbor, MI) ; Brahmasandra; Sundaresh; (Ann Arbor,
MI) ; Kusner; Michael T.; (Ann Arbor, MI) |
|
Applicant: |
Name |
City |
State |
Country |
Type |
NeuMoDx Molecular, Inc. |
Ann Arbor |
MI |
US |
|
|
Family ID: |
53264221 |
Appl. No.: |
15/996949 |
Filed: |
June 4, 2018 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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15413735 |
Jan 24, 2017 |
10010888 |
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15996949 |
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14613616 |
Feb 4, 2015 |
9604213 |
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15413735 |
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13766359 |
Feb 13, 2013 |
9050594 |
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14613616 |
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62065500 |
Oct 17, 2014 |
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61667606 |
Jul 3, 2012 |
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61598240 |
Feb 13, 2012 |
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Current U.S.
Class: |
1/1 |
Current CPC
Class: |
B01L 2400/0406 20130101;
B01L 2400/0633 20130101; B01L 3/50851 20130101; B01L 3/50855
20130101; B01L 2200/10 20130101; B01L 2300/0681 20130101; B01L
2300/0867 20130101; B01L 2300/0887 20130101; B01L 2400/043
20130101; B01L 3/502738 20130101; B01L 2300/168 20130101; B01L
2400/0622 20130101; B01L 2400/0655 20130101; B01L 7/525 20130101;
C12Q 1/686 20130101; B01L 2400/0694 20130101; B01L 2200/0684
20130101; B01L 2200/0689 20130101; B01L 2300/087 20130101; B01L
3/502761 20130101; B01L 2300/044 20130101; B01L 2300/1822 20130101;
B01L 2300/1827 20130101; G01N 35/0098 20130101; B01L 2200/0668
20130101; B01L 2400/0481 20130101; B01L 2400/0487 20130101; B01L
2300/0627 20130101; B01L 2300/0816 20130101; B01L 7/52
20130101 |
International
Class: |
B01L 3/00 20060101
B01L003/00; B01L 7/00 20060101 B01L007/00; C12Q 1/686 20060101
C12Q001/686; G01N 35/00 20060101 G01N035/00 |
Claims
1. A system for processing nucleic acid content of a sample in
cooperation with a cartridge comprising a heating region defined at
a first surface, and a magnet housing region defined by a second
surface in thermal communication with the first surface, the system
comprising: a molecular diagnostic module comprising: a cartridge
platform comprising a magnet receiving slot and operable to receive
the cartridge and align the second surface of the cartridge with
the magnet receiving slot, a heater operable to transmit heat to
the heating region of the cartridge at the first surface, and a
magnet coupled to a magnet heating element that heats the magnet,
the magnet operable to pass through the magnet receiving slot into
the magnet housing region, thereby facilitating separation of a
nucleic acid volume from the sample and heating the sample in
cooperation with the heater.
Description
CROSS-REFERENCE TO RELATED APPLICATIONS
[0001] This application is a is a continuation of U.S. application
Ser. No. 15/413,735, filed 24 Jan. 2017, which is a continuation of
U.S. application Ser. No. 14/613,616, filed 4 Feb. 2015, which is a
continuation-in-part application of U.S. application Ser. No.
13/766,359 filed on 13 Feb. 2013, which claims the benefit of U.S.
Provisional Application Ser. No. 61/667,606, filed on 3 Jul. 2012,
and U.S. Provisional Application Ser. No. 61/598,240, filed on 13
Feb. 2012, which are incorporated herein in their entirety by this
reference. This application is also related to U.S. application
Ser. No. 13/765,996, which is incorporated herein in its entirety
by this reference.
[0002] This application is a is a continuation of U.S. application
Ser. No. 15/413,735, filed 24 Jan. 2017, which is a continuation of
U.S. application Ser. No. 14/613,616, filed 4 Feb. 2015, which also
claims the benefit of U.S. Provisional Application Ser. No.
62/065,500, both of which are incorporated herein in their entirety
by this reference.
TECHNICAL FIELD
[0003] This invention relates generally to the molecular
diagnostics field, and more specifically to an improved system and
method for processing and detecting nucleic acids.
BACKGROUND
[0004] Molecular diagnostics is a clinical laboratory discipline
that has developed rapidly during the last 25 years. It originated
from basic biochemistry and molecular biology research procedures,
but now has become an independent discipline focused on routine
analysis of nucleic acids (NA), including deoxyribonucleic acid
(DNA) and ribonucleic acid (RNA) for diagnostic use in healthcare
and other fields involving analysis of nucleic acids. Molecular
diagnostic analysis of biological samples can include the detection
of one or more nucleic acid materials present in the specimen. The
particular analysis performed may be qualitative and/or
quantitative. Methods of analysis typically involve isolation,
purification, and amplification of nucleic acid materials, and
polymerase chain reaction (PCR) is a common technique used to
amplify nucleic acids. Often, a nucleic acid sample to be analyzed
is obtained in insufficient quantity, quality, and/or purity,
hindering a robust implementation of a diagnostic technique.
Current sample processing methods and molecular diagnostic
techniques are often labor/time intensive, low throughput, and
expensive, and systems of analysis are insufficient. Furthermore,
methods of isolation, processing, and amplification are specific to
certain sample matrices and/or nucleic acid types and not
applicable across common sample and nucleic acid types.
[0005] Due to these and other deficiencies of current molecular
diagnostic systems and methods, there is thus a need for an
improved system and method for processing and detecting nucleic
acids. This invention provides such a system and method.
BRIEF DESCRIPTION OF THE FIGURES
[0006] FIGS. 1A-1B depict an embodiment of a system for processing
and detecting nucleic acids;
[0007] FIGS. 2A-2B depict an embodiment of elements, and a top view
of an embodiment of a system worktable, respectively, of an
embodiment of a system for processing and detecting nucleic
acids;
[0008] FIGS. 3A-3B depict an embodiment of a capture plate for
combining a sample with magnetic beads;
[0009] FIG. 4 depicts an embodiment of a capture plate module to
facilitate lysis of a biological sample and combination of the
biological sample with magnetic beads;
[0010] FIGS. 5A-5B depict an alternative embodiment of a capture
plate;
[0011] FIGS. 6A-6B depict embodiments of a molecular diagnostic
module for processing and detecting nucleic acids;
[0012] FIGS. 7A-7E depict a sequence of operations performed by
elements of an embodiment of a molecular diagnostic module;
[0013] FIG. 8 depicts an embodiment of a microfluidic cartridge and
an embodiment of a cartridge platform;
[0014] FIGS. 9A-9B depict configurations of a linear actuator of an
embodiment of a molecular diagnostic module;
[0015] FIGS. 9C-9E depict configurations of a portion of an
embodiment of a microfluidic cartridge and a molecular diagnostic
module;
[0016] FIGS. 10A-10B depict elements of an embodiment of a valve
actuation subsystem of a molecular diagnostic module;
[0017] FIGS. 11A-11C depict an embodiment of a valve actuation
subsystem of a molecular diagnostic module;
[0018] FIGS. 12A-12D depict elements of an embodiment of an optical
subsystem of a molecular diagnostic module;
[0019] FIG. 13 depicts a side view of an alternative embodiment of
a molecular diagnostic module for processing and detecting nucleic
acids;
[0020] FIGS. 14A-14C depict an embodiment of a fluid handling
system of a system for processing and detecting nucleic acids;
[0021] FIG. 15 depicts embodiments of elements of the fluid
handling system;
[0022] FIGS. 16A-16C are schematics depicting example methods for
processing and detecting nucleic acids;
[0023] FIGS. 17A-17B show embodiments of consumables and reagents
used in a system for processing and detecting nucleic acids;
[0024] FIGS. 18A-18B depict an embodiment of an assay strip to
facilitate analysis of a sample containing nucleic acids;
[0025] FIG. 19 depicts an embodiment of an assay strip holder;
[0026] FIG. 20 depicts an embodiment of an assay strip carrier;
[0027] FIGS. 21A-21B show alternative embodiments of assay strip
holders and assay strips, respectively;
[0028] FIG. 22 shows an embodiment of a filter to facilitate
processing and detecting of nucleic acids;
[0029] FIG. 23 shows an embodiment of a filter holder to facilitate
processing and detecting of nucleic acids; and
[0030] FIGS. 24A-24D depict embodiments of a method for processing
and detecting nucleic acids.
DESCRIPTION OF THE PREFERRED EMBODIMENTS
[0031] The following description of preferred embodiments of the
invention is not intended to limit the invention to these preferred
embodiments, but rather to enable any person skilled in the art to
make and use this invention.
1. System for Processing and Detecting Nucleic Acids
[0032] As shown in FIGS. 1A-1B and 7A, an embodiment of a system
100 for processing and detecting nucleic acids comprises: a capture
plate no configured to facilitate binding of nucleic acids within a
biological sample to a set of magnetic beads 119; a molecular
diagnostic module 130 comprising a microfluidic cartridge receiving
module 140, heating and cooling subsystem 150, a magnet 160, a
valve actuation subsystem 170, an optical subsystem 180; and an
assay strip 190 configured to facilitate mixing of molecular
diagnostic reagents with a nucleic acid volume. Other embodiments
of the system 100 may further comprise at least one of a capture
plate module 120 configured to support the capture plate 110; a
filter 200 and filter holder 205 to facilitate sample preparation;
a microfluidic cartridge 210 configured to facilitate sample
processing; an assay strip holder 230; an assay strip carrier 240;
a liquid handling system 250 configured to facilitate gas and fluid
delivery to different elements of the system 100; a processor
configured to analyze data resulting from a run of the system 100;
and a user interface configured to allow a user to interact with
the system 100. The system 100 thus functions to receive biological
samples containing nucleic acids (i.e., impure nucleic acid
samples), separate nucleic acids from the biological samples, and
analyze nucleic acid samples according to at least one molecular
diagnostic protocol (e.g., PCR). Preferably, the system 100 is a
walkaway system by which a user loads a set of biological samples
containing nucleic acids, and receives a set of data resulting from
a molecular diagnostic protocol without any further sample
manipulation by the user. Alternatively, the system 100 facilitates
aspects of sample preparation for a molecular diagnostic protocol,
with some sample manipulation performed by the user.
[0033] In one example workflow of the system 100, a liquid handling
system 250 aspirates a set of biological samples containing nucleic
acids (i.e., impure nucleic acid samples), and dispenses the set of
biological samples into a capture plate 110 to be lysed and
combined with magnetic beads (containing a proprietary affinity
coating to bind the nucleic acids to the magnetic beads) by a
capture plate module 120. The liquid handling system 250 then
aspirates substantially all of each sample of the set of lysed
biological samples combined with magnetic beads (i.e., set of
magnetic bead-samples) from the capture plate 110, and dispenses
the set of magnetic bead-samples into a microfluidic cartridge 210,
aligned within a cartridge receiving module 140 of a molecular
diagnostic module 130, and configured to be manipulated by the
molecular diagnostic module 130. A heating and cooling subsystem
150, a magnet 160, and a valve actuation subsystem 170 of the
molecular diagnostic module 130 then facilitate separation of a set
of nucleic acids from the magnetic bead-samples, as the liquid
handling system 250 dispenses wash solutions, release solutions,
and/or air at appropriate stages. The liquid handling system 250
then aspirates the set of nucleic acids from the microfluidic
cartridge 210 contained within the molecular diagnostic module 130,
combines the set of nucleic acids with a set of molecular
diagnostic reagents using an assay strip 190, and dispenses the set
of nucleic acids combined with the set of molecular diagnostic
reagents (i.e., set of nucleic acid-reagent mixtures) into the
microfluidic cartridge 210 within the molecular diagnostic module
130. The detection chamber heaters 157, optical subsystem 180 and
valve actuation subsystem 170 of the molecular diagnostic module
130 then facilitate analysis of the set of nucleic acid-reagent
mixtures by a processor configured to display information on a user
interface.
[0034] As stated, the above workflow is just one example workflow
of the system 100, and other workflows of the system 100 and
methods of processing and detecting nucleic acid samples are
further described in Section 2 below. A detailed description of
elements of an embodiment of the system 100 are described in
sections 1.1-1.6 below.
1.1 System--Capture Plate and Capture Plate Module
[0035] As shown in FIGS. 3A and 3B, the capture plate 110 comprises
a capture plate substrate 111 comprising a set of wells 112 and a
puncturable foil seal 115, and functions to facilitate binding of
nucleic acids within a biological sample to a set of magnetic beads
119. Preferably, the entire capture plate 110 is configured to be a
consumable (i.e., disposable), such that each well of the capture
plate 110 can only be used once yet the remaining unused wells can
be used during additional runs of the system 100. Alternatively, at
least a portion of the capture plate 110 is configured to be
reusable, such that additional mixing or reagent additions can be
performed and portions of the capture plate 110 may be used for
multiple runs of the system 100. In one variation of the capture
plate 110, the capture plate substrate 111 is reusable, while the
puncturable foil seal 115 is disposable and replaced after each run
of the system 100.
[0036] The capture plate substrate 111 is configured such that the
capture plate 110 is capable of resting on a flat surface, can be
stacked with another capture plate 110, and also can be manipulated
with industry standard instrument components for handling of
microtiter plates. The capture plate substrate also functions to
define the set of wells 112 and to couple to the puncturable foil
seal 115. The capture plate substrate 111 is preferably composed of
a PCR-compatible polymer that can be heat processed to couple to
the puncturable foil seal 115, but can alternatively be composed of
any appropriate material that can contain a fluid and be bonded to
the puncturable foil seal 115.
[0037] The set of wells 112 of the capture plate substrate 111
function to receive at least one biological sample which contain or
are suspected of potentially containing nucleic acids, and to
facilitate combination of the biological sample with a set of
magnetic beads 119. Preferably, the wells 113 are each configured
to accommodate not only a biological sample, but also to facilitate
mixing of the biological sample with a set of magnetic beads 119
(e.g., using a pipettor, the liquid handling system 250 or other
apparatus), which preferably are preloaded in wells 112, or
alternatively may be added by an operator. Preferably, the wells
are also deeper than they are wide to allow a significant number of
wells 112 (e.g., 24) with clinically relevant sample volumes, and
evenly spaced to facilitate aspiration, delivery, and/or mixing of
multiple biological samples (e.g., with a multi-tip pipettor).
Alternatively, the wells are wider than they are deep to facilitate
larger devices for mixing the biological samples with the magnetic
beads 119. Each well 113 of the set of wells 112 also preferably
has a conically shaped bottom region, as shown in FIG. 3A, to
facilitate complete aspiration of a fluid from a well.
Alternatively, each well 113 may not have a conically shaped bottom
region. Additionally, in the orientation shown in FIG. 3A, the tops
of each well 113 in the set of wells 112 preferably form raised
edges protruding from the capture plate substrate 111, in order to
facilitate sealing of each well 113 by the puncturable foil seal
115. Alternatively, the tops of each well 113 in the set of wells
112 may not form raised edges protruding from the capture plate
substrate in. The magnetic beads are preferably polymer beads,
precoupled with a ligand for binding to a nucleic acid, and
comprising a superparagmagnetic component. Additionally, the
magnetic beads may be treated to be positively charged. However,
the magnetic beads may alternatively be any appropriate magnetic
beads (e.g., magnetic, parmagnetic, or superparamagnetic)
configured to facilitate biomagnetic separation.
[0038] Each quantity of magnetic beads 119 may be accompanied by
lysing reagents (e.g. proteinase K) and a sample process control
comprising nucleic acid sequences for DNA and RNA, which function
to lyse biological samples and to provide a mechanism by which
sample process controls may be later detected to verify processing
fidelity and assay accuracy. The sample process control comprising
nucleic acid sequences for DNA and RNA allows one version of the
capture plate to facilitate assays involving DNA and RNA detection.
Preferably, the quantity of magnetic beads 119, lysing reagents,
and sample process controls is dried within each well to improve
shelf life; however, the quantity of magnetic beads 119, lysing
reagents, and sample process controls may alternatively be in
liquid form.
[0039] The puncturable foil seal 115 functions to isolate each well
113 of the set of wells 112, prevent contamination of the contents
of each of the set of wells 112, protect the magnetic beads 119 and
other reagents stored in wells 112 from degradation, and provide
information identifying the capture plate 110. The puncturable foil
seal 115 preferably seals each well 113 of the capture plate 110,
and is configured to be punctured by an external element (e.g., by
a pipette tip), such that each well is sealed prior to being
punctured. In one variation, the puncturable foil seal 115 also
forms a seal around an element that punctures it, and in another
variation, the puncturable foil seal 115 does not form a seal
around an element that punctures it, in order to prevent airlock.
The puncturable foil seal 115 is also preferably labeled with
identifying information including at least one of manufacturer
information, capture plate contents, the lot of the contents, an
expiry date, and a unique electronic tag (e.g., barcode or QR code)
providing more information. Preferably, the puncturable foil seal
115 does not extend beyond the footprint of the capture plate 110,
but alternatively, the puncturable foil seal 115 may be any
appropriate size and/or include protruding features (e.g., tabs)
that facilitate handling of the capture plate.
[0040] In one variation, the capture plate 110 may be prepackaged
at least with magnetic beads 119, such that each well 113 in the
set of wells 112 is prepackaged with a set of magnetic beads 119
defined by a specific quantity or concentration of magnetic beads.
The set of wells 112 may then be sealed by the puncturable foil
seal 115, which is configured to be punctured by an external
element that delivers volumes of biological samples to be mixed
with the magnetic beads 119. In another variation, the capture
plate 110 may not be prepackaged with magnetic beads 119, but the
wells 113 of the capture plate may still be sealed with a
puncturable foil seal 115. In this variation, the puncturable foil
seal 115 is configured to be punctured by at least one external
element, for co-delivery of biological samples and magnetic beads
intended to be combined.
[0041] A variation of the capture plate 110' may further comprise a
slotted rubber membrane 116, as shown in FIGS. 5A and 5B,
configured to provide access through the puncturable foil seal 115
to the set of wells 112. The slotted rubber membrane 116 thus
functions to prevent or reduce splashing, evaporation, and/or
aerosolization of contents of the set of wells 112. Preferably, the
slotted rubber membrane 116 comprises slots that are self-sealing
and centered over wells of the set of wells 112, and further does
not extend beyond the footprint of the capture plate 110.
Alternatively, the slots of the slotted rubber membrane 116 may not
be self-sealing, and/or the slotted rubber membrane 116 may be any
appropriate size and comprise features that extend beyond the
footprint of the capture plate 110.
[0042] In a specific example, the capture plate 110 comprises 24
wells 113 with an 18 mm center-to-center pitch, each well having a
volumetric capacity of 2 mL, and is compliant with Society for
Laboratory Automation and Screening (SLAS) standards. Each well 113
of the capture plate 110 in the specific example is also
prepackaged with a specified quantity of magnetic beads 119, and
comprises a protruding top edge that is heat sealed to a
puncturable foil seal. In addition, each well 113 also contains
other reagents beneficial for processing and monitoring the sample,
including proteinase K and one or more specific nucleic acid stands
designed to serve as a process control. The specific example of the
capture plate 110 can thus combine two groups of 12 biological
samples with magnetic beads. The capture plate no in the specific
example is produced by injection molding, has a footprint of 127.75
mm.times.85.5 mm, and is composed of a PCR-compatible polypropylene
based polymer with a high vapor barrier.
[0043] An embodiment of the system 100 may further comprise a
capture plate module 120, as shown in FIG. 4, which functions to
receive, support, and heat a capture plate 110. The capture plate
module 120 preferably comprises a thermally conducting substrate
121 configured to cradle a capture plate 110, a capture plate
heater 123, a capture plate receiving module 125, and a capture
plate electronics module 127. Preferably, the capture plate module
120 functions to facilitate lysis of a biological sample deposited
into a well 113 of the capture plate, and to facilitate binding of
nucleic acids (i.e., within a lysed biological sample) to a
quantity of magnetic beads 119 within a well 113 of the capture
plate 110. In a specific example, the capture plate module 120 has
dimensions of 108 mm.times.156 mm.times.45 mm and is configured to
rest on a flat surface.
[0044] The thermally conducting substrate 121 is configured to
cradle and support the capture plate 110, and functions to conduct
heat to the set of wells 112 of the capture plate 110. Preferably,
the thermally conducting substrate 121 is also configured to
reversibly couple to the capture plate 110, and comprises a set of
indentations 122 that encircle each well 113 in the set of wells
112. In one variation, the indentations 122 completely conform to
the external surface of each well 113 of the capture plate 110, but
in another variation, the indentations 122 may encircle a portion
of each well 113 of the capture plate 110. Additionally, the
indentations 122 are preferably thermally conducting in order to
conduct heat to the set of wells 112, and portions of the thermally
conducting substrate 121 aside from the indentations 122 are
composed of non-conducting, rigid material. Alternatively, the
entire thermally conducting substrate 121 may be composed of a
material that is thermally conducting.
[0045] The capture plate heater 123 is preferably coupled to the
thermally conducting substrate 121, and functions to transfer heat,
through the thermally conducting substrate 121, to a well 113 of
the capture plate 110. The capture plate heater 123 preferably
conforms to at least a portion of an indentation 122 of the
thermally conducting substrate 121, to facilitate heat transfer
through the indentation 122 to an individual well 113 of the
capture plate 110. In this variation, the capture plate heater 123
is one of a set of capture plate heaters 124, wherein each capture
plate heater 123 in the set of capture plate heaters 124 transfers
heat to an individual well 113 of the set of wells 112 of the
capture plate 110. Alternatively, the capture plate heater 123 may
conform to portions of multiple indentations 122 of the thermally
conducting substrate 121, such that the capture plate heater 123 is
configured to transfer heat to multiple wells 113 of the capture
plate 110. Preferably, the capture plate heater 123 is a resistance
heater, but alternatively, the capture plate heater 123 may be a
Peltier or any appropriate heater configured to transfer heat to
the capture plate 110. The capture plate heater 123 may also
further couple to a heat sink.
[0046] The capture plate receiving module 125 comprises a capture
plate actuation system 126 that functions to couple the capture
plate module 120 to a capture plate 110. As shown in FIG. 4, the
capture plate actuation system 126 comprises a structural support
with hinged grips 128 and at least one capture plate module
actuator 129. The capture plate module actuator 129 is preferably a
push-type solenoid with a spring return, but may alternatively be
any appropriate linear actuator, such as a hydraulic actuator. The
structural support with hinged grips 128 preferably couples to the
capture plate heater 123 and houses the capture plate module
actuator 129, such that, in a first configuration, actuation of the
capture plate module actuator 129 outwardly displaces the hinged
grips (allowing the capture plate module 120 to receive a capture
plate no), and in a second configuration, actuation of the capture
plate module actuator 129 inwardly displaces the hinged grips
(allowing the capture plate module 120 to couple to the capture
plate 110). The structural support with hinged grips 128 may
further comprise a textured and/or high-friction surface configured
to grip a capture plate 110, but alternatively may not comprise a
textured and/or high-friction surface.
[0047] The capture plate electronics module 127 is coupled to the
capture plate heater 123 and the capture plate actuation system
126, and functions to enable control of the capture plate heater
123 and the capture plate actuation system 126. Preferably, the
capture plate electronics module 127 modulates an output of the
capture plate heater 123, in order to controllably heat at least
one well 113 of the capture plate 110. Additionally, the capture
plate electronics module 127 preferably modulates the capture plate
actuation system 126, in order to controllably couple the capture
plate module 120 to a capture plate 110. Preferably, the capture
plate electronics module 127 is coupled to an external power
supply, such that the capture plate module 120 does not include an
integrated power supply; however, in alternative embodiments, the
capture plate electronics module 127 may be coupled to a power
supply integrated with the capture plate module 120.
1.2 System--Molecular Diagnostic Module
[0048] As shown in FIGS. 6A and 6B, an embodiment of the molecular
diagnostic module 130 of the system 100 includes a cartridge
receiving module 140, a heating and cooling subsystem 150, a magnet
160, a valve actuation subsystem 170, and an optical subsystem 180,
and functions to manipulate a microfluidic cartridge 210 for
processing of a biological sample containing nucleic acids. The
molecular diagnostic module 130 is preferably configured to operate
in parallel with at least one other molecular diagnostic module
130, such that multiple microfluidic cartridges 210 containing
biological samples may be processed simultaneously. In a first
variation, the molecular diagnostic module 130 is configured to be
stackable with another molecular diagnostic module 130 in a manner
that enables access to a microfluidic cartridge 210 within each
molecular diagnostic module 130; an example of the first variation
is shown in FIG. 6B, where the molecular diagnostic modules 130 are
stacked in a staggered configuration. In the first variation, each
molecular diagnostic module 130 may further comprise locking pins
or other appropriate mechanisms to couple the stacked molecular
diagnostic modules 130 together. In another variation, the
molecular diagnostic module 130 may not be configured to stack with
another molecular diagnostic module, such that the molecular
diagnostic modules 130 are configured to rest side-by-side on the
same plane. Elements of an embodiment of the molecular diagnostic
module 130 are further described in sections 1.2.1 to 1.2.5
below.
1.2.1 Molecular Diagnostic Module--Cartridge Receiving Module
[0049] As shown in FIG. 9A, the cartridge receiving module 140 of
the molecular diagnostic module 130 comprises a cartridge platform
141 including a cartridge loading guiderail 142, a cartridge stop
143, a magnet receiving slot 144, and a set of valve actuation
slots 145; a linear actuator 146 configured to displace a
microfluidic cartridge 210 resting on the cartridge platform 141;
and a set of springs 148 coupled to the cartridge platform 141. The
cartridge receiving module 140 thus functions to receive, align,
and compress a microfluidic cartridge 210 for processing of a
biological sample according to a molecular diagnostic assay
protocol. As shown in FIGS. 7A-7C, the cartridge platform 141 is
preferably configured to receive a microfluidic cartridge 210 along
a cartridge loading guiderail 142 until it reaches a cartridge stop
143, and be vertically displaced by the linear actuator 146, which
places a biasing force against the set of springs 148 coupled to
the cartridge platform 141. The magnet receiving slot 144 and the
set of valve actuation slots 145 provide access, by a magnet 160
and a valve actuation subsystem 170, to the microfluidic cartridge
210, as the microfluidic cartridge is vertically displaced by the
linear actuator 146.
[0050] The cartridge platform 141 includes a cartridge loading
guiderail 142, a cartridge stop 143, a magnet receiving slot 144,
and a set of valve actuation slots 145, and functions to receive
and align a microfluidic cartridge 210, while providing access to
the microfluidic cartridge 210 by a magnet 160 and a valve
actuation subsystem 170. As shown in FIG. 8, an embodiment of the
cartridge platform 141 includes a pair of parallel cartridge
loading guiderails 142, initiating at a pair of inwardly tapering
protrusions configured to guide a microfluidic cartridge toward the
pair of parallel cartridge loading guiderails 142, and spanning two
short edges of the cartridge platform 141. The embodiment of the
cartridge platform 141 also includes a cartridge stop 143
comprising a vertical tab oriented perpendicular to the cartridge
loading guiderails 142, and spanning a long edge of the cartridge
platform. Preferably, the cartridge loading guiderails 142 and the
cartridge stop 143 are configured such that a microfluidic
cartridge 210 slides between the cartridge loading guiderails 142
and hits the cartridge stop 143 to signal proper alignment.
Alternatively, the cartridge loading guiderails 142 and the
cartridge stop 143 may be configured such that a microfluidic
cartridge slides over or along the cartridge loading guiderails
142, after which the cartridge stop 143 couples to a portion of the
microfluidic cartridge 210 to ensure proper alignment of the
microfluidic cartridge. Additional variations of the cartridge
loading guiderails 142 and the cartridge stop 143 may be used to
enable reception and alignment of a microfluidic cartridge 210 by
the molecular diagnostic module 130, and are known by those skilled
in the art.
[0051] The embodiment of the cartridge platform 141 shown in FIG. 8
also includes a set of valve actuation slots 145, oriented
perpendicular to the parallel cartridge loading guiderails 142 and
configured to provide access to a valve actuation subsystem 170,
and a magnet receiving slot 144 located among the set of valve
actuation slots 145. Preferably, the magnet receiving slot 144 and
the set of valve actuation slots 145 substantially span a long
dimension of the cartridge platform 141, as shown in FIG. 8, and
are configured to correspond to locations on a microfluidic
cartridge 210 requiring a magnetic field and/or valving to enable
processing of a biological sample and nucleic acid detection once
the microfluidic cartridge 210 has been aligned within the
molecular diagnostic module 130. Thus, alternative configurations
of the magnet receiving slot 144 and the set of valve actuation
slots 145 may accommodate other cartridges with alternative regions
requiring magnetic fields and/or valving to enable other protocols.
In one alternative embodiment, the magnet receiving slot 144 and
the set of valve actuation slots may comprise one continuous void
of the cartridge platform 141, such that the cartridge platform 141
supports a microfluidic cartridge 210 along the periphery of the
microfluidic cartridge 210, but forms a continuous void under a
majority of the footprint of the microfluidic cartridge 210.
[0052] The linear actuator 146 functions to linearly displace a
microfluidic cartridge 210 resting on the cartridge platform 141,
in order to compress the microfluidic cartridge 210 and position
the microfluidic cartridge 210 between a cartridge heater 153 and
an optical subsystem 180 on one side of the microfluidic cartridge
210, and a magnet 160 and detection chamber heaters 157 on another
side of the microfluidic cartridge 210. The linear actuator 146
also functions to provide a sufficient counterforce to the valve
actuation subsystem 170 such that a microfluidic cartridge 210
within the molecular diagnostic module 130 remains properly
situated upon manipulation by the valve actuation subsystem 170.
The linear actuator 146 further functions to move a nozzle 149
coupled to the liquid handling system 250, in order to couple the
liquid handling system 250 to a fluid port 222 of the microfluidic
cartridge 210. In the orientation of the molecular diagnostic
module 130 shown in FIGS. 7B and 7B, the linear actuator 146 is
preferably coupled to a portion of the heating and cooling
subsystem 150 a portion of the optical subsystem 180, and the
nozzle 149, and vertically displaces the cartridge heater 153, the
optical subsystem 180, and the nozzle 149 to position the cartridge
heater 153, 180 and the nozzle 149 over the microfluidic cartridge
210. The vertical displacement also allows the microfluidic
cartridge 210 to receive a magnet 160, which provides a magnetic
field to facilitate a subset of a molecular diagnostic protocol,
and detection chamber heaters 157, which allows amplification of
nucleic acids for molecular diagnostic protocols requiring heating
and cooling of the nucleic acid (e.g. PCR). Preferably, the linear
actuator 146 is a scissor jack actuator configured to apply
substantially uniform pressure over all occlusion positions of a
microfluidic cartridge 210 aligned within the molecular diagnostic
module 130, and to operate in at least two configurations. In a
retracted configuration 146a, as shown in FIG. 9A, the scissor jack
actuator has not linearly displaced the cartridge platform 141, and
in an extended configuration 146b, as shown in FIG. 9B, the scissor
jack actuator has linearly displaced the microfluidic cartridge 210
to position the microfluidic cartridge 210 between the subsystems
153, and 180, and the magnet 160 and detection chamber heaters 157.
Additionally, the extended configuration 146b of the scissor jack
actuator is configured to couple the nozzle 149 to a fluid port 222
of the microfluidic cartridge 210, such that the liquid handling
system 250 can deliver solutions and gases for processing of
biological samples. The linear actuator 146 may alternatively be
any appropriate linear actuator, such as a hydraulic, pneumatic, or
motor-driven linear actuator, configured to linearly displace a
microfluidic cartridge within the molecular diagnostic module
130.
[0053] As shown in FIGS. 7B, 7C, and 8, a set of springs 148 is
coupled to the cartridge platform 141 and functions to provide a
counteracting force against the linear actuator 146 as the linear
actuator 146 displaces a microfluidic cartridge 210 resting on the
cartridge platform 141. The set of springs 148 thus allows the
cartridge platform 141 to return to a position that allows the
microfluidic cartridge 210 to be loaded and unloaded from the
molecular diagnostic module 130 when the linear actuator 146 is in
a retracted configuration 146b, as shown in FIG. 7B. Preferably, in
the orientation shown in FIG. 7B, the set of springs 148 is located
at peripheral regions of the bottom side of the cartridge platform
141, such that the set of springs 148 does not interfere with the
magnet or the valve actuation subsystem 170. Alternatively, the set
of springs 148 may be located at any appropriate position to
provide a counteracting force against the linear actuator 146. In a
specific example shown in FIG. 6A, the set of springs 148 comprises
four springs located near corners of the bottom side of the
cartridge platform 141, but in other variations, the set of springs
148 may comprise any appropriate number of springs. Each spring of
the set of springs 148 is also preferably housed within a guide to
prevent deviations from linear vertical motions (in the orientation
shown in FIG. 7B); however, each spring in the set of springs 148
may alternatively not be housed within a guide. In an alternative
embodiment of the molecular diagnostic module 130, the set of
springs 148 may altogether be replaced by a second linear actuator
configured to linearly displace a microfluidic cartridge 210,
resting on the cartridge platform 141, in a direction opposite to
the displacements enforced by the linear actuator 146, and/or by
any other suitable element (e.g., elastomeric element) configured
to provide a biasing force against a microfluidic cartridge 210 at
the cartridge platform 141.
[0054] Similarly, the nozzle 149, the heating and cooling subsystem
150, the cartridge heater 153, and the magnet 160 are preferably
coupled to springs, such that springs are positioned between
elements 149, 150, 153, and 160, and substrates that elements 149,
150, 153, and 160 are mounted to. Alternatively an elastomeric
material is preferably positioned between elements 149, 150, 153,
and 160, and substrates that elements 149, 150, 153, and 160 are
mounted to. The springs and/or elastomeric material function to
provide proper functioning and alignment of subsystems of the
molecular diagnostic module 130 as the linear actuator 146 is
extended or retracted, contributing to reliability and a reduction
in stack up tolerance risk. The springs and/or elastomeric material
further function to allow more pressure to be applied to occlusion
positions of a microfluidic cartridge 210 aligned within the
molecular diagnostic module 130, and an appropriate pressure to be
applied to elements 149, 150, 153 and 160 of the molecular
diagnostic module 130. Thus, proper contact is maintained between
elements 149, 150, 153, and 160, and a microfluidic cartridge 210
being manipulated by the molecular diagnostic module. These
elements are described in further detail below.
1.2.2 Molecular Diagnostic Module--Heating/Cooling Subsystem and
Magnet
[0055] The heating and cooling subsystem 150 of the molecular
diagnostic module 130 comprises a cartridge heater 153, a fan 155,
and a set of detection chamber heaters 157 and functions to
controllably heat portions of a microfluidic cartridge 210 for
processing of a biological sample containing nucleic acids
according to a molecular diagnostic protocol. In the orientation of
an embodiment of the molecular diagnostic module 130 shown in FIGS.
7A-7C, the cartridge heater 153 is preferably coupled to the linear
actuator 146 of the cartridge receiving module 140 and configured
to span a central region of a microfluidic cartridge 210 aligned
within the molecular diagnostic module 130, the fan 155 is located
at a back wall of the cartridge receiving module 140, and the set
of detection chamber heaters 157 is located inferior to a set of
detection chambers 213 of the microfluidic cartridge 210. In
alternative embodiments of the molecular diagnostic module 130, the
heating and cooling subsystem 150 may have any appropriate
alternative configuration that provides controlled heating and
cooling to a microfluidic cartridge within the molecular diagnostic
module 130.
[0056] The cartridge heater 153 functions to transfer heat to a
heating region 224 of a microfluidic cartridge 210, for inducing a
pH shift to release bound nucleic acids from magnetic beads within
the heating region 224. The cartridge heater 153 is preferably a
plate-shaped heater configured to transfer heat to the microfluidic
cartridge 210 only from one side of the cartridge heater 153, such
that heat flows through one face of the plate-shaped heater to the
microfluidic cartridge 210. In a specific example, the cartridge
heater 153 is a silicon wafer etched to be conductive and form a
resistance heater. In the preferred variation, the cartridge heater
153 is either flip-chip bonded (i.e., soldered to back side of a
circuit board), or wire bonded to a circuit board, and then coupled
using linear bearings and springs to a plate coupled to the linear
actuator 146. The preferred variation allows independent control of
12 independent channels, corresponding to 12 different pathways for
sample processing. In another variation, heating through one face
is accomplished using a plate-shaped resistance heater that has one
exposed face and thermal insulation covering all other faces, and
in yet another variation heating through one face is accomplished
using a Peltier heater. In a variation of the cartridge heater 153
using a Peltier heater, the cartridge heater 153 comprises a
thermoelectric material, and produces different temperatures on
opposite faces of the cartridge heater 153 in response to a voltage
difference placed across the thermoelectric material. Thus, when a
current flows through the Peltier heater, one face of the Peltier
heater lowers in temperature, and another face of the Peltier
heater increases in temperature. Alternative variations of the
cartridge heater 153 can be used to appropriately transfer heat to
a heating region 224 of the microfluidic cartridge 210.
[0057] Preferably, the cartridge heater 153 is configured to
linearly translate with the linear actuator 146 of the cartridge
receiving module 140, in order to align with a heating region 224
spanning a central portion of a microfluidic cartridge 210 aligned
within the molecular diagnostic module 130. In one variation, the
cartridge heater 153 is preferably fixed relative to the linear
actuator 146 such that (in the orientation shown in FIGS. 7B-7C),
the cartridge heater 153 can only move vertically with the linear
actuator. In an alternative variation, the cartridge heater 153 may
additionally be configured to translate laterally with a horizontal
plane (in the orientation shown in FIGS. 7B-7C), such that the
cartridge heater 153 can translate in at least two perpendicular
coordinate planes. In this alternative variation, the cartridge
heater 153 can be configured to sweep across a surface of a
microfluidic cartridge 210 aligned within the molecular diagnostic
module 130, or to translate in response to motion of the
microfluidic cartridge 210, such that the position of the cartridge
heater 153 relative to a heating region 224 of the microfluidic
cartridge 210 is always fixed.
[0058] The fan 155 functions to modulate heat control within the
molecular diagnostic module 130, by enabling heat transfer from
warm objects within the molecular diagnostic module 130 to cooler
air external to the molecular diagnostic module 130. In the
orientation shown in FIG. 6A, the fan 155 is preferably located at
a back face of the molecular diagnostic module 130, such heat
within the molecular diagnostic module 130 is transferred out of
the back face of the molecular diagnostic module 130 to cooler air
external to the molecular diagnostic module. In a specific
embodiment, the molecular diagnostic module 130 comprises four fans
155 located at the back face of the molecular diagnostic module
130; however, in alternative embodiments the molecular diagnostic
module 130 may comprise any appropriate number of fans located at
any appropriate position of the molecular diagnostic module 130. In
one variation, the fan 155 may be passive and driven solely by
convection currents resulting from motion of hot air within the
molecular diagnostic module to cooler air outside of the molecular
diagnostic module; however, in alternative variations, the fan 155
may be motor-driven and configured to actively cool internal
components of the molecular diagnostic module 130 if molecular
diagnostic module elements exceed a certain threshold
temperature.
[0059] The set of detection chamber heaters 157 functions to
individually heat detection chambers of a set of detection chambers
213 within a microfluidic cartridge 210. Each detection chamber
heater in the set of detection chamber heaters 157 is preferably
configured to heat one side of one detection chamber in the set of
detection chambers 213, and is preferably located such that the
extended configuration 146b of the linear actuator 146 of the
cartridge receiving module 140 puts a detection chamber in
proximity to a detection chamber heater. As mentioned above, the
set of detection chamber heaters 157 is preferably coupled to
springs or an elastomeric layer to ensure direct contact between
the set of detection chamber heaters and a set of detection
chambers, without compressively damaging the set of detection
chamber heater 157. Preferably, each detection chamber heater is
configured to contact a surface of a detection chamber in the
extended configuration 146b of the linear actuator 146; however,
each detection chamber heater may be further configured to couple
to a detection chamber in the extended configuration 146b of the
linear actuator 146. In a first variation, the set of detection
chamber heaters 157 comprises silicon chip heaters flip chipped to
one surface of a flexible printed circuit board, with a set of
springs coupled to an opposite surface of the flexible printed
circuit board, such that each spring in the set of springs aligns
with a detection chamber heater. In the first variation, contact
between each detection chamber heater and a detection chamber is
thus maintained by a biasing force provided by an individual spring
through the flexible printed circuit board. In a second variation,
the set of detection chamber heaters 157 comprises silicon chip
heaters flip chipped to one surface of a rigid printed circuit
board, with a set of springs coupled to an opposite surface of the
rigid printed circuit board. In the second variation, the set of
springs thus function to collectively transfer a force through the
rigid printed circuit board to maintain contact between the set of
detection chamber heaters and a set of detection chambers.
Preferably, each detection chamber heater in the set of detection
chamber heaters 157 is configured to contact and heat a bottom
surface of a detection chamber (in the orientation shown in FIG.
7B); however, each detection chamber heater may alternatively be
configured to contact and heat both a top and a bottom surface of a
detection chamber. Additionally, each detection chamber heater
preferably corresponds to a specific detection chamber of the set
of detection chambers 213 and functions to individually heat the
specific detection chamber; however, alternatively, each detection
chamber heater may be configured to heat multiple detection
chambers in the set of detection chambers 213. Preferably, all
detection chamber heaters in the set of detection chamber heaters
157 are identical; however, the set of detection chamber heaters
157 may alternatively not comprise identical detection chamber
heaters.
[0060] In one variation, each detection chamber heater in the set
of detection chamber heaters 157 comprises a donut-shaped heater,
configured to encircle a surface of a detection chamber. The
donut-shaped heater may further include a conducting mesh
configured to allow detection through the heater while still
allowing efficient heat transfer to the detection chamber. In an
alternative variation, each detection chamber heater in the set of
detection chamber heaters 157 may include a plate-shaped Peltier
heater, similar to Peltier cartridge heater 153 described above. In
this alternative variation, each detection chamber heater is thus
configured to heat one side of a detection chamber through one face
of the detection chamber heater. In one specific example, the
molecular diagnostic module 130 comprises 12 diced silicon wafers
with conductive channels flip chipped to 12 detection chambers,
providing resistive heating to each of the 12 detection chambers.
In another specific example, the molecular diagnostic module 130
comprises a 12 Peltier detection chamber heaters configured to heat
12 detection chambers of a microfluidic cartridge 210 aligned
within the molecular diagnostic module 130. In other alternative
variations, each detection chamber heater may comprise any
appropriate heater configured to individually heat a detection
chamber.
[0061] In some variations, reflection from the set of detection
chamber heaters 157 can interfere with light transmitted to
photodetectors of the optical subsystem 180 (e.g., light emitted
from the set of biological samples, light transmitted through
filters of an optical subsystem), especially in configurations
wherein the set of detection chambers 213 of a microfluidic
cartridge 210 are positioned between detection chamber heaters 157
and optical elements of an optical subsystem 180. In these
variations, the set of detection chamber heaters 157 can include
elements that reduce or eliminate reflection from the set of
detection chamber heaters 157, thereby facilitating analysis of the
set of biological samples. In one variation, the set of detection
chamber heaters 157 can include or be coupled to non-reflective
coatings at surfaces of the set of detection chamber heaters 157
upon which light from the optical subsystem 180 impinges. In a
specific example, the non-reflective coating can comprise a
high-temperature paint (e.g., dark paint, flat paint) that absorbs
and/or diffuses light from the optical subsystem 180, while
facilitating heat transfer to a set of detection chambers 213 of a
microfluidic cartridge 210. In another variation, the set of
detection chamber heaters can be in misalignment with
photodetectors of the optical subsystem 180, such that reflection
does not interfere with light transmitted to the photodetectors of
the optical subsystem 180. In one example, the set of detection
chamber heaters can be configured to heat a set of detection
chambers 213 from a first direction, and the optical subsystem 180
can be configured to receive light from the set of detection
chambers 213 from a second direction (e.g., a direction
non-parallel to the first direction), such that reflection from the
detection chamber heaters 157 does not cause interference. In still
other variations, the set of detection chamber heaters 157 can
include any other suitable elements (e.g., coatings, layers, etc.)
and/or be configured in any other suitable manner that eliminates,
prevents, or mitigates reflection from the set of detection chamber
heaters 157 from interfering with light transmitted to
photodetectors of the optical subsystem 180.
[0062] The magnet 160 of the molecular diagnostic module 130
functions to provide a magnetic field for isolation and extraction
of nucleic acids bound to magnetic beads within a microfluidic
cartridge 210, aligned within the molecular diagnostic module 130.
Preferably, the magnet 160 is fixed within the molecular diagnostic
module 130, such that the extended configuration 146b of the linear
actuator 146 allows the magnet 160 to pass through the magnet
receiving slot 144 of the cartridge receiving module 140 and into a
magnet housing region 218 of the microfluidic cartridge 210. In an
example, as shown in FIGS. 7A-7C, the magnet 160 is a rectangular
prism-shaped magnet 160 fixed under the cartridge platform 141, and
configured to pass through the cartridge platform 141, into a
magnet housing region 218 located at a surface of the microfluidic
cartridge 210 directly opposing the heating region 224 of the
microfluidic cartridge 210. As such, in some variations, a
projection of the heating region 224 onto a plane at least
partially overlaps with a projection of the magnet housing region
218 onto the plane. However, the magnet housing region 218 and the
heating region 224 of the microfluidic cartridge 210 can be in
thermal communication in any other suitable manner. Preferably, the
magnet 160 is one of multiple magnets (e.g., 2-3 magnets) lined up
in parallel, such that each of the fluidic pathways of a
microfluidic cartridge housing the magnets is exposed to a
multiplied magnetic flux (e.g., two or three times as much magnetic
flux), and a multiplied opportunity to capture magnetic beads.
Alternatively, the magnet 160 is a single magnet configured to
expose a set of fluidic pathways to a magnetic field. Preferably,
the magnet 160 or group of multiple magnets is coupled to a magnet
holder within the molecular diagnostic module 130. Additionally,
the magnet holder is preferably composed of an insulating material,
such that the magnet holder does not interfere with proper
functioning of the cartridge heater 153. Alternatively, the magnet
holder may not be composed of an insulating material.
[0063] In one variation, the magnet 160 or group of multiple
magnets comprises a permanent magnet, composed of a magnetized
material (e.g., a ferromagnet) providing a substantially fixed
magnetic field. In an alternative variation, the magnet 160 or
group of multiple magnets comprises an electromagnet configured to
provide a modifiable magnetic field, such that the intensity of the
magnetic field can be adjusted, the polarity of the magnetic field
can be reversed, and the magnetic field can be substantially
removed upon removal of a current flowing within the electromagnet.
Preferably, the magnet 160 or group of magnets is also fixed
relative to the molecular diagnostic module 130; however, the
magnet 160 or group of magnets may alternatively be configured to
translate vertically (in the orientation shown in FIG. 7B), such
that the magnet 160 or group of magnets can extend into and retract
from the magnet receiving slot 144 of the cartridge platform 141
and the magnet housing region 218 of the microfluidic cartridge
210. Additionally, the magnet 160 or group of magnets preferably
rides on linear bearings and/or springs (or an elastomeric
material) to ensure proper contact with a microfluidic cartridge in
an extended configuration 146b of the linear actuator 146, in a
manner that allows most of force from the linear actuator 146 to
translate to full occlusion of a subset of the set of occlusion
positions (i.e., without leakage).
[0064] In some variations, wherein a magnet housing region 218 of
the microfluidic cartridge 210 is located at a surface of the
microfluidic cartridge 210 directly opposing the heating region 224
of the microfluidic cartridge 210, all or a subset of the magnet(s)
16o can be heated, such that the magnet(s) do not provide a heat
sink at surfaces of the microfluidic cartridge 210 opposing the
cartridge heater 153 and/or any other portion of the microfluidic
cartridge 210 intended to have a desired heated state (e.g., a
portion of the microfluidic cartridge proximal the set of detection
chamber heaters 157). Preferably, heating of the magnet(s) 160 of
the molecular diagnostic module is performed in a manner that does
disrupt alignment of magnetic domains (e.g., for a permanent
magnet), such that a magnetic field provided by the magnet(s) 160
does not diminish in strength. As such, a magnet 160 of the
molecular diagnostic module 130 is preferably heated to a
temperature less than its Curie point, or can additionally or
alternatively comprise a magnetic material with a sufficiently high
Curie point (e.g., a Curie point characterized by a higher
temperature than temperatures required for processing of samples at
the microfluidic cartridge 210). In one example, the magnet(s) 160
can thus be configured to be heated to one or more temperatures in
synchronization with temperatures of the cartridge heater 153, in
order to further increase uniformity of heating through the
microfluidic cartridge (e.g., from a heating region 224 to a
magnet-housing region). The magnet 160 can, however, be any other
suitable magnet (e.g., permanent magnet, electromagnet) that is not
disrupted by heating within the range of temperatures required for
processing of samples at the microfluidic cartridge 210.
Furthermore, the magnet(s) 160 of the molecular diagnostic module
130 can be configured to be heated with any suitable temperature
output, such that the magnet(s) facilitate generation of any
suitable heating profile (e.g., non-uniform heating profile,
uniform heating profile, etc.) through the microfluidic cartridge
210.
[0065] In one variation, the molecular diagnostic module 130 can
comprise at least one magnet 160 coupled to a magnet heating
element, such that the magnet heating element heats the magnet 160
to a desired state. In one example of this variation, the molecular
diagnostic module 130 can comprise a set of magnets 260, wherein
each magnet 160 of the set of magnets 260 is coupled to a magnet
heating element 261 at least at one surface of the magnet. As such,
the magnet heating element can be coupled to a surface of the
magnet 160, can wrap about multiple surfaces of the magnet, can be
at least partially embedded in the magnet, and/or can be coupled to
the magnet in any other suitable manner. In one specific example,
as shown in FIG. 9E, each magnet 160 of the set of magnets 260 is
separated from an adjacent magnet by a magnet heating element 261,
and in another specific example, each magnet 160 of the set of
magnets has a magnet heating element coupled to a distal end of the
magnet 160, wherein the distal end of the magnet 160 is configured
to interface with the magnet housing region 218 of the microfluidic
cartridge 210. The magnet(s) 160 and the magnet heating element(s)
261 of the microfluidic cartridge 210 can, however, be configured
in any other suitable manner.
[0066] Additionally or alternatively, the molecular diagnostic
module 130 can be configured with an insulation gap between the
magnet(s) 160 and a surface of the microfluidic cartridge 210
proximal the magnet housing region 218, such that the magnet(s) 160
do not interfere with heating of the microfluidic cartridge 210.
The insulation gap can be an air gap within the system or can
additionally or alternatively comprise any other suitable
insulating layer situated between the magnet(s) 160 and the surface
of the microfluidic cartridge 210 opposing the cartridge heater
153.
[0067] In any of the above embodiments and variations of the
magnet(s), the magnet(s) are preferably configured to span a
substantial portion of a capture segment 263 (e.g., an s-shaped
capture segment with a characteristic width) of the microfluidic
cartridge 210, by way of the magnet housing region 218, wherein the
capture segment is a portion of a fluidic pathway configured to
facilitate capture of target particles bound to magnetic particles.
As such, the magnet is preferably substantially wide in order to
span a majority of the capture segment and provide a desired
gradient of magnetic strength at the capture segment, by way of the
magnet housing region 218 of the microfluidic cartridge 210.
Additionally, the strength of the magnet(s) can be adjusted to
prevent clogging within the capture segment, for instance, by
adjusting morphology, composition, and/or any other suitable
characteristic of the magnet(s). In one specific example, the
magnet is wide enough to span a majority, but not all, of an
s-shaped capture segment 263 of a microfluidic cartridge, by
crossing the s-shaped capture segment in an orientation
perpendicular to a flow direction through the s-shaped capture
segment 163, as shown in FIGS. 9C-9E; however, the magnet(s) 160
can alternatively be configured in any other suitable manner.
[0068] Alternative configurations and/or compositions of the magnet
160 may also be appropriate in facilitating isolation and
extraction of nucleic acids bound to magnetic beads within the
microfluidic cartridge 210.
1.2.3 Molecular Diagnostic Module--Valve Actuation Subsystem
[0069] As shown in FIGS. 10A-11C, the valve actuation subsystem 170
of the molecular diagnostic module 130 comprises a set of pins 172
configured to translate linearly within a pin housing 175, by
sliding a cam card 177 laterally over the pins 172. The valve
actuation subsystem 170 functions to provide a biasing force to
deform an object in contact with the set of pins 172. In a
configuration wherein a microfluidic cartridge 210 is aligned
within the molecular diagnostic module 130, the valve actuation
subsystem 170 thus functions to occlude a fluidic pathway 220 of
the microfluidic cartridge 210 at a set of occlusion positions 226,
to control flow of a biological sample containing nucleic acids,
reagents and/or air through the microfluidic cartridge 210. In an
embodiment of the molecular diagnostic module shown in FIGS. 7D-7E,
the set of pins 172 and the pin housing are located directly under
the microfluidic cartridge 210, such that the set of pins can
access the microfluidic cartridge 210 through the valve actuation
accommodating slots 145 of the cartridge platform 141. The cam card
177 in the embodiment is positioned under the set of pins and is
coupled to a linear cam card actuator 178 configured to laterally
displace the cam card 177 to vertically displace pins of the set of
pins 172. Preferably, as shown in FIG. 11A, the cam card 177 rests
on a low friction surface configured to facilitate lateral
displacement of the cam card 177; however, the cam card 177 may
alternatively rest on a bed of ball bearings to facilitate lateral
displacement of the cam card 177, or may rest on any feature that
allows the cam card 177 to be laterally displaced by the linear cam
card actuator 178.
[0070] The cam card 177, as shown in FIGS. 7D and 11A, includes a
set of hills 176 and valleys 179, and functions to transform linear
motion in one plane to vertical motion in another plane. In one
variation, the cam card 177 is coupled to a linear actuator and
contacts the ends of pins in a set of pins 172, such that when a
hill 176 of the cam card 177 passes under a pin, the pin is in a
raised configuration 177a, and when a valley 179 of the cam card
177 passes under a pin, the pin is in a lowered configuration 177b.
The hills 176 and valleys 179 of the cam card 177 are preferably in
a set configuration, as shown in FIG. 11B, such that lateral motion
of the cam card 177 to a set position raises a fixed subset of the
set of pins 172. In this manner, lateral movement of the cam card
177 to different positions of a set of positions consistently
raises different subsets of the set of pins 172 to occlude
different portions of a fluidic pathway 220 of a microfluidic
cartridge 210 in contact with the set of pins 172. Thus, portions
of a fluidic pathway 220 may be selectively occluded and opened to
facilitate processing of a biological sample according to any
appropriate tissue, cellular, or molecular diagnostic assay
protocol. In one variation, the cam card is configured to be
laterally displaced in two coordinate directions within a plane
(e.g., by x-y linear actuators), and in another variation, the cam
card is configured to be laterally displaced in only one coordinate
direction within a plane (e.g., by a single linear actuator). In a
specific example, the hills 176 of the cam card 177 are raised 1 mm
above the valleys 179 of the cam card 177, the hills 176 and
valleys 179 each have a 2 mm wide plateau region, and a hill 176
region slopes down to a valley region 179 at a fixed angle over a 2
mm length. In the specific example, the cam card 177 is driven by a
Firgelli linear actuator. Alternative variations may include any
appropriate configurations and geometries of a cam card with hills
176 and valleys 179, driven by any appropriate actuator.
[0071] In alternative embodiments of the valve actuation subsystem
170, the cam card 177 may be a cam card wheel comprising a set of
hills 176 and valleys 179 on a cylindrical surface, and configured
to convert rotary motion to linear (i.e., vertical) motion of the
set of pins 172. The cam card wheel may be configured to contact
ends of pins in the set of pins 172, and may be coupled to a motor
shaft and driven by a motor. In other alternative embodiments of
the valve actuation subsystem 170, the cam card 177 may altogether
be replaced by a set of cams, each configured to individually
rotate about an axis. In these alternative embodiments, rotating
subsets of the set of cams raises corresponding subsets of the set
of pins, and occludes specific portions of a fluidic pathway 220 of
a microfluidic cartridge 210 in contact with the set of pins
172.
[0072] The set of pins 172 functions to selectively occlude
portions of a fluidic pathway 220 of a microfluidic cartridge 210
at least at subsets of a set of occlusion positions 226. The pins
of the set of pins 172 are preferably cylindrical and, in the
orientation shown in FIG. 11A, configured to slide over a cam card
177 and within a pin housing 175. Each pin in the set of pins 172
preferably also includes a first spring 173 that functions to
provide a counteracting force to restore a pin to a lowered
configuration 177b; however, each pin in the set of pins 172 may
alternative not include a first spring 173, and rely solely on
gravity to return to a lowered configuration 177b. Preferably, as
shown in FIG. 11C, each pin is also composed of two parts separated
by a second spring, which functions to allow sufficient force to
fully occlude a microfluidic channel but prevents forces from being
generated that could damage the pin, microfluidic cartridge and/or
cam card. Each pin also preferably comprises a first region 171
configured to slide within the pin housing 175, and a second region
174 configured to exit the pin housing 175. The second region 174
is preferably of a smaller dimension than the first region 171,
such that each pin is constrained by the pin housing 175 to be
raised by a limited amount. Alternatively, the first region 171 and
the second region 174 may have any appropriate configuration to
facilitate raising and lowering of a pin by a fixed amount. In a
specific example, the valve actuation subsystem 170 comprises 12
sets of pins 172 configured to selectively occlude 12 fluidic
pathways 212 of a microfluidic cartridge 210 aligned within the
molecular diagnostic module; however, other embodiments may
comprise any appropriate number of sets of pins 172.
[0073] In the orientation shown in FIG. 11A, each pin in the set of
pins 172 preferably has a circular cross section and round ends,
configured to facilitate sliding within a pin housing 175, sliding
over a cam card 177 surface, and occlusion of a fluidic pathway
220. Alternatively, each pin may comprise any appropriate
cross-sectional geometry (e.g., rectangular) and/or end shape
(e.g., flat or pointed) to facilitate occlusion of a fluidic
pathway 220. Preferably, the surface of each pin in the set of pins
172 is composed of a low-friction material to facilitate sliding
motions (i.e., over a cam card 177 or within a pin housing 175);
however, each pin may alternatively be coated with a lubricant
configured to facilitate sliding motions.
[0074] The pin housing 175 functions to constrain and guide the
motion of each pin in the set of pins 172, as the cam card 177
slides under the set of pins 172. Preferably, the pin housing 175
comprises a set of pin housing channels 169 configured to surround
at least one pin in the set of pins 172. In one variation, each pin
in the set of pins 172 is surrounded by an individual channel of
the set of pin housing channels 169; however, in another variation
a channel of the set of pin housing channels 169 may be configured
to surround multiple pins in the set of pins 172. In an example
shown in FIGS. 7D-7E and 11A, the pin housing is located under the
cartridge platform 141, such that the set of pin housing channels
169 is aligned with the set of valve actuation accommodating slots
145, to provide access, by the set of pins 172, to a microfluidic
cartridge 210 aligned on the cartridge platform 141. In the
example, the pin housing 175 thus constrains the set of pins 172,
such that each pin can only move linearly in a vertical direction.
Each pin housing channel preferably has a constricted region 168
configured to limit the motion of a pin within a pin channel;
however, each pin housing channel may alternatively not include a
constricted region. Preferably, surfaces of the pin housing 175
contacting the set of pins 172 are composed of a low friction
material to facilitate sliding of a pin within a pin housing
channel; however, surfaces of the pin housing 175 contacting the
set of pins 172 may alternatively be coated with a lubricant
configured to facilitate sliding motions. Other variations of the
pin housing 175 and the set of pins 172 may include no additional
provisions to facilitate sliding of a pin within a pin housing
channel.
[0075] In some embodiments of the molecular diagnostic module 130,
the valve actuation subsystem 170 can be configured in any other
suitable manner to facilitate actuation of a set of pins 172 to
occlude a microfluidic cartridge 210 at a set of occlusion
positions 226. In one embodiment, the valve actuation subsystem 170
can be an embodiment of the valve actuation subsystem described in
U.S. application Ser. No. 14/229,396, entitled "System and Method
for Processing Biological Samples" and filed on 28 Mar. 2014, which
is incorporated herein in its entirety by this reference.
1.2.4 Molecular Diagnostic Module--Optical Subsystem
[0076] As shown in FIGS. 12A-12D, the optical subsystem 180 of the
molecular diagnostic module 130 comprises a set of light emitting
diodes (LEDs) 181, a set of excitation filters 182 configured to
transmit light from the set of LEDs 181, a set of dichroic mirrors
183 configured to reflect light from the set of excitation filters
182 toward a set of apertures 185 configured to transmit light
toward a set of nucleic acid samples, a set of emission filters 186
configured to receive and transmit light emitted by the set of
nucleic acid samples, and a set of photodetectors 187 configured to
facilitate analysis of light received through the set of emission
filters 186. The optical subsystem 180 may further comprise a set
of lenses 184 configured to focus light onto the set of nucleic
acid samples. The optical subsystem 180 thus functions to transmit
light at excitation wavelengths toward a set of nucleic acid
samples and to receive light at emission wavelengths from a set of
nucleic acid samples. Preferably, the optical subsystem 180 is
coupled to an optical subsystem actuator 188 configured to
laterally displace and align the optical subsystem 180 relative to
a set of nucleic acid samples at a set of detection chambers 213 of
the microfluidic cartridge 210, and is further coupled to a linear
actuator 146 of the cartridge receiving module 140 to position the
optical subsystem 180 closer to the set of nucleic acid samples.
Alternatively, the optical subsystem 180 may not be coupled to a
linear actuator 146 of the cartridge receiving module 140, and may
only be configured to translate laterally in one direction. In a
specific example, the optical subsystem 180 comprises a set of 12
apertures, a set of 12 lenses, a set of 12 dichroic mirrors, a set
of 12 excitation filters, a set of 12 LEDs, a set of 12 emission
filters, and a set of 12 photodetectors. In a variation of the
specific example, each set of 12 optical subsystem 180 elements is
configured as two sets of 6 apertures, two sets of 6 lenses, two
sets of 6 dichroic mirrors, two sets of 6 excitation filters (e.g.,
6 excitation filters spanning different wavelengths of light), two
sets of 6 LEDs, two sets of 6 emission filters (e.g., 6 emission
filters spanning different wavelengths of light, and two sets of 6
photodetectors, such that the optical subsystem 180 includes two
sets of identical optical subsystem units, each comprising 6 of
each element. The optical subsystem 180 can, however, comprise any
suitable number of identical units, in order to increase throughput
in the system 100 by decreasing the amount of time it takes to
analyze multiple biological samples. For instance, multiple
identical units can enable analyses to be performed using a unit of
the optical subsystem 180, at a desired point during sample
processing (e.g., a point at which a biological sample in a
microfluidic cartridge is being heated). In variations of the
optical subsystem 180 comprising multiple units, each unit can be
configured to move independently of the other units, or can
additionally or alternatively be configured to move with the other
units in a non-independent manner. For instance, one optical
subsystem unit can be configured to move along a first axis (e.g.,
a horizontal axis) independently of the other optical subsystem
unit(s), but can be configured to move along a second axis (e.g., a
vertical axis) non-independently of the other optical subsystem
unit(s).
[0077] In the specific examples, as shown in FIG. 7A-7E, the
optical subsystem 180 is located within the molecular diagnostic
module 130 and coupled to the linear actuator 146 of the cartridge
receiving module 140, such that, in the extended configuration 146b
of the linear actuator 146, the optical subsystem 180 can be
positioned closer to a microfluidic cartridge 210 aligned within
the molecular diagnostic module. Conversely in the specific
example, the optical subsystem 180 is positioned away from the
microfluidic cartridge 210 in the retracted configuration 146a of
the linear actuator 146. In the specific example, the optical
subsystem 180 is further coupled to an optical subsystem actuator
188 configured to laterally displace the optical subsystem 180
relative to the microfluidic cartridge 210, such that the optical
subsystem 180 can be aligned with a set of detection chambers 213
of the microfluidic cartridge 210.
[0078] Preferably, the set of LEDs 181 are not all identical but
rather chosen to efficiently produce a certain band of wavelengths
of light, such that light from the set of LEDs 181 can be filtered
to appropriate narrow wavelengths for analysis of nucleic acid
samples. Alternatively, all LEDs in the set of LEDs 181 may be
identical, and produce white light comprising all wavelengths of
visible light that is filtered to produce the desired wavelength,
in which case the LEDs may be stationary. Preferably, the set of
LEDs 181 includes phosphor-based LEDs, but the set of LEDs 181 may
alternatively include any LEDs configured to provide light of the
desired range of wavelengths. The LEDs of the set of LEDs 181 are
preferably configured to emit light of wavelengths corresponding to
at least one of the set of excitation filters 182, the set of
dichroic mirrors 183, and the set of emission filters 186.
[0079] The set of excitation filters 182 is configured to align
with the set of LEDs 181 in the optical subsystem 180, and
functions to transmit light at excitation wavelengths toward the
set of dichroic mirrors 183 of the optical subsystem 180.
Preferably, the set of excitation filters 182 are not identical
excitation filters, but rather chosen to transmit the different
desired ranges of excitation wavelengths. Alternatively, all
excitation filters of the set of excitation filters 182 are
identical, and configured to transmit light having a fixed range of
excitation wavelengths. In one variation, the set of excitation
filters 182 includes band pass filters, configured to transmit
light between two bounding wavelengths, in another variation, the
set of excitation filters 182 includes short pass filters
configured to transmit light below a certain wavelength, and in yet
another variation, the set of excitation filters 182 includes long
pass filters configured to transmit light above a certain
wavelength. Preferably, the set of excitation filters 182 is
interchangeable, such that individual excitation filters may be
interchanged to provide different excitation wavelengths of light;
however, the set of excitation filters 182 may alternatively be
fixed, such that the optical subsystem 180 is only configured to
transmit a fixed range of excitation wavelengths.
[0080] The set of dichroic mirrors 183 is configured to align with
the set of excitation filters 182, and functions to receive and
reflect light from the set of excitation filters 182 toward the
detection chamber, such that light having a range of excitation
wavelengths may be focused, through a set of apertures, onto a set
of nucleic acid samples. The set of dichroic mirrors 183 also
functions to receive and transmit light from a set of emission
filters 186 toward a set of photodetectors 187, which is described
in more detail below. All dichroic mirrors in the set of dichroic
mirrors 183 are preferably identical in orientation relative to the
set of excitation filters 182 and the set of emission filters 186,
and configured to reflect and transmit the appropriate wavelengths
of light for the given LED. Alternatively, the set of dichroic
mirrors 183 may include identical dichroic mirrors, with regard to
orientation, light transmission, and light reflection. In a
specific example, in the orientation shown in FIG. 12A, the set of
excitation filters 182 is oriented perpendicular to the set of
emission filters 186, with the set of dichroic mirrors 183
bisecting an angle between two planes formed by the faces of the
set of excitation filters 182 and the set of emission filters 186.
In the specific example, light from the set of excitation filters
is thus substantially reflected at a 90.degree. angle toward the
set of apertures 185, and light from the set of emission filters
186 passes in a substantially straight direction through the set of
dichroic mirrors 183 toward the set of photodetectors 187. Other
variations of the set of dichroic mirrors 183 may include any
configuration of dichroic mirrors, excitation filters, and/or
emission filters that enable transmission of light of excitation
wavelengths toward a set of nucleic acid samples, and transmission
of light from the set of nucleic acid samples toward a set of
photodetectors 187.
[0081] In one embodiment, the optical subsystem may further include
a set of lenses 184 configured to align with the set of dichroic
mirrors 183, which functions to focus light, from the set of
excitation filters 182 and reflected off of the set of dichroic
mirrors 183, onto a set of nucleic acid samples configured to emit
light in response to the light from the set of excitation filters
182. All lenses in the set of lenses 184 are preferably identical
in orientation relative to the set of dichroic mirrors and in
dimension; however, the set of lenses 184 may alternatively
comprise non-identical lenses, such that light passing through
different lenses of the set of lenses 184 is focused differently on
different nucleic acid samples. In a specific example, in the
orientation shown in FIG. 12A, the faces of the set of lenses 184
are oriented perpendicular to the faces of the set of excitation
filters 182, to account for light reflection by the set of dichroic
mirrors 183 at a 90.degree. angle. In the specific example, the set
of lenses also includes identical 1/4'' high numerical aperture
lenses. In other variations, the set of lenses 184 may be oriented
in any appropriate configuration for focusing light from the set of
dichroic mirrors 183 onto a set of nucleic acid samples, and may
include lenses of any appropriate specification (i.e., numerical
aperture).
[0082] The set of apertures 185 is located on an aperture substrate
189 and configured to align with the set of lenses 184, and
functions to allow focused light from the set of lenses 184 to pass
through to the set of nucleic acid samples. The aperture substrate
189 is preferably coupled to the linear actuator 146 of the
cartridge receiving module 140, which allows the optical subsystem
180 to linearly translate and be positioned near and away from a
microfluidic cartridge 210 aligned within the molecular diagnostic
module 130. Alternatively, the aperture substrate 189 may not be
coupled to the linear actuator 146 of the cartridge receiving
module 140. Preferably, all apertures 185 in the set of apertures
185 are identical, and configured to allow identical light profiles
to be focused, through the set of lenses 184, onto a set of nucleic
acid samples. Alternatively, the set of apertures 185 may not
include identical apertures. In one variation, each aperture in the
set of apertures 185 may be individually adjustable, in order to
provide individually modifiable aperture dimensions (e.g., width,
length, or diameter) to affect light exposure. In an alternative
variation, each aperture in the set of apertures 185 is fixed.
Other variations may include interchangeable aperture substrates
189, such that features of the set of apertures (e.g., aperture
dimensions, number of apertures) may be adjusted by interchanging
aperture substrates 189.
[0083] The set of emission filters 186 is configured to align with
the set of dichroic mirrors, and functions to transmit emission
wavelengths of light from the set of nucleic acid samples, and to
filter out excitation wavelengths of light. Preferably, each
emission filter of the set of emission filters 186 are configured
to transmit light having a fixed range of emission wavelengths,
while blocking light of excitation wavelengths. Alternatively, the
set of emission filters 186 may comprise identical emission
filters, such that individual emission filters of the set of
emission filters 186 are configured to transmit the same ranges of
emission wavelengths. In one variation, the set of emission filters
186 includes band pass filters, configured to transmit light
between two bounding wavelengths, in another variation, the set of
emission filters 186 includes short pass filters configured to
transmit light below a certain wavelength, and in yet another
variation, the set of emission filters 186 includes long pass
filters configured to transmit light above a certain wavelength.
Preferably, the set of emission filters 186 is interchangeable,
such that individual emission filters may be interchanged to
transmit and/or block different wavelengths of light; however, the
set of emission filters 186 may alternatively be fixed, such that
the optical subsystem 180 is only configured to transmit a fixed
range of emission wavelengths.
[0084] The set of photodetectors 187 is configured to align with
the set of emission filters 186, and functions to receive light
from the set emission filters to facilitate analysis of the set of
nucleic acid samples. All photodetectors in the set of
photodetectors 187 are preferably identical; however, the set of
photodetectors 187 may alternatively include non-identical
photodetectors. Preferably, the set of photodetectors 187 includes
photodiodes comprising a photoelectric material configured to
convert electromagnetic energy into electrical signals; however,
the set of photodetectors 187 may alternatively comprise any
appropriate photodetectors for facilitating analysis of biological
samples, as is known by those skilled in the art.
[0085] The optical subsystem actuator 188 is coupled to the optical
subsystem 180, and functions to laterally translate the optical
subsystem 180 relative to a set of nucleic acid samples being
analyzed. Preferably, the optical subsystem actuator 188 is a
linear actuator configured to translate the optical subsystem 180
in one dimension; however, the optical subsystem actuator 188 may
alternatively be an actuator configured to translate the optical
subsystem 180 in more than one dimension. In a specific example, as
shown in FIGS. 7A-7D and 12D, the optical subsystem actuator 188 is
configured to translate the optical subsystem 180 laterally in a
horizontal plane, to align the optical subsystem 180 with a set of
detection chambers 213 of a microfluidic cartridge 210 within the
molecular diagnostic module 130. In another example, the optical
subsystem may be configured as a disc revolving around an axis with
the LEDs and photodetectors stationary and the disc containing the
filters. In other variations, the optical subsystem actuator 188
may be configured in any appropriate manner to facilitate alignment
of the optical subsystem 180 relative to a set of nucleic acid
samples being analyzed.
[0086] In some variations, wherein reflection from the set of
detection chamber heaters 157 and/or any other element of the
system interferes with light emitted directly from biological
samples at the microfluidic cartridge 210 or light transmitted
through the set of excitation filters 182, the optical system can
be configured to filter out undesired reflected light, by way of
any one or more of: the set of excitation filters 182, the set of
dichroic mirrors 183, the set of emission filters 186, and any
other suitable element configured to reduce or remove interference
caused by undesired reflected light.
[0087] The optical subsystem 180 can, however, comprise any other
suitable element(s) and/or be configured in any other suitable
manner to facilitate analysis of a set of biological samples.
1.2.5 Molecular Diagnostic Module--Alternative Embodiments and
Variations
[0088] As described above, alternative embodiments of the molecular
diagnostic module 130 and alternative variations of subsystems and
elements of the molecular diagnostic module 130 may be configured
to process a biological sample containing nucleic acids, isolate
nucleic acids from the biological sample, and detect nucleic acids.
An example of an alternative embodiment of a molecular diagnostic
module 130, as shown in FIG. 13, includes a cartridge receiving
module 140', a heating and cooling subsystem 150', a magnet 160', a
valve actuation subsystem 170', and an optical subsystem 180', and
functions to manipulate an alternative microfluidic cartridge 210'
for processing of biological samples containing nucleic acids.
Other alternative embodiments of the molecular diagnostic module
130'' may be configured to receive alternative microfluidic
cartridges 210'', for processing of biological samples containing
nucleic acids.
1.3 System--Assay Strip
[0089] As shown in FIGS. 18A and 18B, the assay strip 190 comprises
an assay strip substrate 191 comprising a set of wells 192, and
typically a puncturable foil seal 195, and functions to facilitate
combination of a set of nucleic acid samples with a set of
molecular diagnostic reagents for amplification and/or detection of
a nucleic acid sequence or sequences. Preferably, the entire assay
strip 190 is configured to be a consumable (i.e., disposable), such
that the assay strip 190 can be used during multiple runs of the
system 100, then the assay strip 190 is disposed of once all of the
wells 192, containing unitized reagents for a single test or group
of tests, is exhausted. Alternatively, at least a portion of the
assay strip 190 is configured to be reusable, such that wells may
be reloaded with reagents and reused with the system 100. In one
variation of the assay strip 190, the assay strip substrate 191 is
reusable, while the puncturable foil seal 195 is disposable and
replaced after each run of the system 100. In another variation,
the reusable assay strip substrate 191 does not require a
puncturable foil seal 195, such that reagents specific to a certain
nucleic acid sequences may be deposited into open wells of the
assay strip substrate 191 by a user.
[0090] The assay strip substrate 191 is configured such that the
assay strip 190 is capable of resting on a flat surface, and
functions to define the set of wells 192 and to couple to the
puncturable foil seal 195. The assay strip substrate 191 is
preferably configured to be received by a corresponding assay strip
holder 230 configured to hold multiple assay strips 190, but may
alternatively not be configured to couple to an assay strip holder
230. The assay strip substrate 191 is preferably composed of a
PCR-compatible polymer, such as polypropylene, that can be heat
processed to couple to the puncturable foil seal 115, but can
alternatively be composed of any appropriate material that can
contain a fluid and be bonded to the puncturable foil seal 115.
[0091] The set of wells 192 of the assay strip substrate 191
function to receive at least one nucleic acid sample, and to
facilitate combination of the nucleic acid sample with at least one
of a set of molecular diagnostic reagents. The molecular diagnostic
reagents of the set of molecular diagnostic reagents preferably
comprise reagents configured to analyze the set of nucleic acid
volumes for markers of at least one of gonorrhea (GC), Chlamydia
(CT), herpes simplex virus (HSV), human immunodeficiency virus
(HIV), human respiratory diseases, vaginal diseases, hepatitis C
virus (HCV), hepatitis B virus (HBV), trichonomas, group B
streptococcus (GBS), factor 2 (FII) gene, and factor five (FV)
gene, but may alternatively comprise reagents used to perform
alternative molecular diagnostic protocols. Preferably, the wells
193 of the assay strip substrate 191 are each configured to
accommodate not only a nucleic acid sample, but also to facilitate
mixing of the nucleic acid sample with at least one of a set of
molecular diagnostic reagents (e.g., using a pipettor or other
apparatus). Additionally, the molecular diagnostic reagents of the
set of molecular diagnostic reagents preferably comprises probes
and primers to detect the sample process controls provided by the
capture plate, in order to verify process fidelity and assay
accuracy. Preferably, the wells 193 are deep enough to facilitate
mixing without splashing, and evenly spaced to facilitate
aspiration, delivery, and/or mixing of multiple biological samples
(e.g., with a multi-tip pipettor). Alternatively, the wells are
wide and shallow to facilitate drying of reagents in the wells to
increase shelf life and larger devices for mixing the nucleic acids
with molecular diagnostic reagents. Each well 193 of the set of
wells 192 also preferably has a rounded bottom region, as shown in
FIG. 18A, to facilitate complete aspiration of a fluid from a well
193; however, each well 193 may alternatively not have a rounded
bottom region. Additionally, the set of wells 192 is preferably
arranged in staggered rows, which functions to facilitate access to
individual wells 193 of the set of wells, to reduce one dimension
of the assay strip 190, and also to prevent cross-contamination of
fluids within the wells due to dripping. Alternatively, the set of
wells 192 may not be arranged in staggered rows.
[0092] The puncturable foil seal 195 functions to protect the
molecular diagnostic reagents stored in wells 112 from degradation,
isolate each well 193 of the set of wells 192, prevent
contamination of the contents of each of the set of wells 192, and
provide information identifying the assay strip 190. The
puncturable foil seal 195 preferably seals each well 193 of the
assay strip 190, and is configured to be punctured by an external
element (e.g., by a pipette tip), such that each well is sealed
prior to being punctured. In one variation, the puncturable foil
seal 195 also forms a seal around an element that punctures it, and
in another variation, the puncturable foil seal 195 does not form a
seal around an element that punctures it, in order to prevent
airlock. The puncturable foil seal 195 is also preferably labeled
with identifying information including at least one of manufacturer
information, assay strip contents, the lot of the contents, an
expiry date, and a unique electronic tag (e.g., barcode or QR code)
providing more information. Preferably, the puncturable foil seal
195 does not extend beyond the footprint of the assay strip 190,
but alternatively, the puncturable foil seal 195 may be any
appropriate size and/or include protruding features (e.g., tabs)
that facilitate handling of the assay strip.
[0093] In one variation, the assay strip 190 may be prepackaged
with a set of molecular diagnostic reagents, such that each well
193 in the set of wells 192 is prepackaged with a quantity of
molecular diagnostic reagents. The set of wells 192 may then be
sealed by the puncturable foil seal 195, which is configured to be
punctured by an external element that delivers volumes of nucleic
acid samples to be combined with the set of molecular diagnostic
reagents. In another variation, the assay strip 190 may not be
prepackaged with a set of molecular diagnostic reagents, and the
wells 193 of the assay strip 190 may not be sealed with a
puncturable foil seal 195. In yet another variation, the system may
comprise an empty assay strip 190 without a puncturable foil seal
195, and an assay strip 190 comprising reagents and a puncturable
foil seal 195, such that a user may add specific reagents to the
empty assay strip to be used in conjunction with the assay strip
comprising reagents. In variations comprising a puncturable foil
seal 195, the puncturable foil seal 115 is configured to be
punctured by at least one external element, for co-delivery of
nucleic acid samples and molecular diagnostic reagents intended to
be combined.
[0094] In a specific example, the assay strip 190 has an 87
mm.times.16 mm footprint and comprises 24 wells 113 arranged in two
staggered rows, with a 9 mm center-to-center pitch between adjacent
wells 193 within each row. Each well 193 of the set of wells has a
capacity of 60 .mu.L to accommodate a volume of a molecular
diagnostic reagent, 20 .mu.L of a sample fluid, and any
displacement caused by a pipette tip (e.g., 100 or 300 .mu.L
pipette tip). Each well 113 of the assay strip 190 in the specific
example is also prepackaged with a quantity of molecular diagnostic
reagents, and comprises a protruding top edge (75 microns high)
that is heat sealed to a puncturable foil seal. The capture plate
110 in the specific example is produced by injection molding, has a
footprint of 127.75 mm.times.85.5 mm, and is composed of a
PCR-compatible polypropylene based polymer with a high vapor
barrier. In the specific embodiment, the vapor barrier is further
increased by depositing a thin metallic layer to the outside of the
assay strip 190.
[0095] As described earlier, the assay strip 190 may be configured
to be received by an assay strip holder 230. The assay strip holder
230 functions to receive and align multiple assay strips 190, such
that a multichannel pipettor or other fluid delivery system may
combine multiple nucleic acid samples with molecular diagnostic
reagents using wells 193 of multiple assay strips 190. In one
variation, the assay strip holder 230 may be configured to contain
Assay strips 190 including reagents for substantially different
molecular diagnostic assays, as shown in FIG. 17B, such that a
single run of the system 100 involves analyzing a set of nucleic
acid samples under different molecular diagnostic assays. In
another variation, the assay strip holder 230 may be configured to
contain assay strips 190 including reagents for identical molecular
diagnostic assays, such that a single run of the system 100
involves analyzing a set of nucleic acid samples under the same
molecular diagnostic assay. Preferably, the assay strip holder 230
is composed of a material that is dishwasher safe and autoclavable,
configured to hold the assay strips 190 in place during handling by
a fluid delivery system (e.g., pipettor), and configured such that
the assay strips 190 avoid protruding over an edge of the assay
strip holder 230, but the assay strip holder 230 is constructed to
facilitate insertion and removal of the assay strips 190 from the
assay strip holder 230.
[0096] In one variation, the assay strip holder 230 is not
configured to facilitate cooling of molecular diagnostic reagents
within the assay strips 190; however, in another variation as shown
in FIG. 21A, the assay strip holder 230 may be further configured
to couple to an aluminum block 235 coupled to a set of Peltier
units 236 configured to facilitate cooling of molecular diagnostic
reagents within the assay strips 190. Additionally, the assay strip
holder 230 may be configured to be received and carried by an assay
strip carrier 240, which, as shown in FIG. 20, functions to
facilitate handling and alignment of multiple assay strip holders
230. In a specific example, as shown in FIG. 19, the assay strip
holder 230 has dimensions of 127.76 mm.times.85.48 mm.times.14.35
mm, complies with American National Standards Institute (ANSI) and
Society for Laboratory Automation and Screening (SLAS) standards,
and is configured to hold six 16-well assay strips for a total of
96 wells 193. In another specific example, as shown in FIG. 21B,
the assay strip holder 230' is configured to hold four assay strips
190', each comprising 24 wells 193' for a total of 96 wells per
assay strip holder 230'. Other combinations of the described
embodiments, variations, and examples of the assay strip 190, assay
strip holder 230, and assay strip carrier 240 may be incorporated
into embodiments of the system 100 for processing and detecting
nucleic acids.
1.4 System--Microfluidic Cartridge
[0097] The microfluidic cartridge 210 functions to receive a set of
magnetic bead-samples, facilitate separation of nucleic acids from
the set of magnetic bead-samples, receive a set of nucleic
acid-reagent samples, and facilitate analysis of nucleic acids from
the set of nucleic acid-reagent samples. In one embodiment, the
microfluidic cartridge 210 comprises a top layer 211 including a
set of sample port-reagent port pairs 212 and a set of detection
chambers 213; an intermediate substrate 214, coupled to the top
layer 211 and partially separated from the top layer 211 by a film
layer 215, configured to form a waste chamber 216; an elastomeric
layer 217 partially situated on the intermediate substrate 214; a
magnet housing region 218 accessible by a magnet 160 providing a
magnetic field; and a set of fluidic pathways 219, each formed by
at least a portion of the top layer 211, a portion of the film
layer 215, and a portion of the elastomeric layer 217. In the
embodiment, the microfluidic cartridge 10 further comprises a
bottom layer 221 coupled to the intermediate substrate 214 and
configured to seal the waste chamber 216. Furthermore, in the
embodiment, the top layer 211 of the microfluidic cartridge 210
further comprises a shared fluid port 222, a vent region 223, and a
heating region 224, such that each fluidic pathway 220 in the set
of fluidic pathways 219 is fluidically coupled to a sample
port-reagent port pair 224, the shared fluid port 222, the waste
chamber 216, and a detection chamber 225, comprises a turnabout
portion 226 configured to pass through the heating region 224 and
the magnetic field, and is configured to pass through the vent
region 223 upstream of the detection chamber 225. Each fluidic
pathway 220 thus functions to receive and facilitate processing of
a sample fluid containing nucleic acids as it passes through
different portions of the fluidic pathway 220.
[0098] The microfluidic cartridge 210 is preferably configured to
be received and manipulated by the molecular diagnostic module 130,
such that the cartridge receiving module 140 of the molecular
diagnostic module 130 receives and aligns the microfluidic
cartridge 210 within the molecular diagnostic module 130, the
heating and cooling subsystem 150 of the molecular diagnostic
module 130 is configured to transfer heat to the heating region 224
of the microfluidic cartridge 210, and the magnet 160 of the
molecular diagnostic module 130 is configured to be received by the
magnet housing region 218 of the microfluidic cartridge 210 to
provide a magnetic field for separation of nucleic acids.
Additionally, the shared fluid port 222 of the microfluidic
cartridge 210 is configured to couple to a nozzle 149 coupled to
the linear actuator 146 of the cartridge receiving module 140, such
that the liquid handling system 250 can deliver fluids and gases
through the shared fluid port 222. The elastomeric layer 217 of the
microfluidic cartridge 210 is also preferably configured to be
occluded at a set of occlusion positions 226 by the valve actuation
subsystem 170 of the molecular diagnostic module, in order to
occlude portions of a fluidic pathway 220 of the microfluidic
cartridge 210 for processing of a set of biological samples. The
optical subsystem 180 of the molecular diagnostic module 130 is
further configured to align with the set of detection chambers 213
of the microfluidic cartridge 210, to facilitate analysis of a set
of nucleic acid samples. The microfluidic cartridge 210 is
preferably the microfluidic cartridge 210 described in U.S.
application Ser. No. 13/765,996, which is incorporated in its
entirety by this reference, but may alternatively be any
appropriate cartridge or substrate configured to receive and
process a set of samples containing nucleic acids.
1.5 System--Fluid Handling System and Filter
[0099] The liquid handling system 250 of the system 100 includes a
liquid handling arm 255 and a syringe pump 265, as shown in FIGS.
14A-14C and functions to deliver biological samples, reagents, and
gases to elements of the system 100. As described in Section 1, an
embodiment of the liquid handling system 250 is configured to
aspirate a set of biological samples containing nucleic acids
(i.e., impure nucleic acid samples), dispense the set of biological
samples into a capture plate 110 to be lysed and combined with
magnetic beads by a capture plate module 120, aspirate the set of
biological samples combined with magnetic beads (i.e., set of
magnetic bead-samples) from the capture plate 110, and dispense the
set of magnetic bead-samples into microfluidic cartridge 210
located in a molecular diagnostic module 130. The embodiment of the
liquid handling system 100 is further configured to facilitate
separation of a set of nucleic acids from the magnetic
bead-samples, by dispensing a wash solution, a release solution,
and/or air into the molecular diagnostic module 130, by the nozzle
149 coupled to the linear actuator 146, at appropriate stages,
aspirate the set of nucleic acids from the molecular diagnostic
module 130, combine the set of nucleic acids with a set of
molecular diagnostic reagents using an assay strip 190, and
dispense the set of nucleic acids combined with the set of
molecular diagnostic reagents (i.e., set of nucleic acid-reagent
mixtures) into the molecular diagnostic module 130 for further
processing and analysis. Other embodiments of the liquid handling
system 250 may be configured to perform alternative molecular
diagnostic assay protocols and/or dispense and aspirate alternative
fluids into and from other elements supporting a molecular
diagnostic protocol.
[0100] The liquid handling arm 255 comprises a gantry 256 and a
multichannel liquid handling head 257, and functions to travel to
different elements of the system 100 for fluid delivery and
aspiration. The liquid handling arm 255 is preferably automated and
configured to move, aspirate, and deliver fluids automatically, but
may alternatively be a semi-automated liquid handling arm 255
configured to perform at least one of moving, aspirating, and
delivering automatically, while another entity, such as a user,
performs the other functions.
[0101] The gantry 256 is coupled to the multichannel liquid
handling head 257, and functions to transport the multichannel
liquid handling head 257 to different elements of the system 100
for fluid delivery and aspiration. Preferably, the gantry 256 is
automated and configured to translate the multichannel liquid
handling head 257 within at least two dimensions, and provides X-Y
positional accuracy of at least 0.5 mm. Additionally, in the
orientation shown in FIG. 14B, the gantry is preferably situated
above the molecular diagnostic module 130, such that the gantry 256
can translate within at least two dimensions without interfering
with other elements of the system 100. Alternatively, the gantry
256 may be any appropriate gantry 256 to facilitate movement of an
end effector within at least two dimensions, as is readily known by
those skilled in the art.
[0102] The multichannel liquid handling head 257 functions to
aspirate fluids from and deliver fluids to different elements of
the system 100. Preferably, the multichannel liquid handling head
257 is a multichannel pipette head; however, the multichannel
liquid handling head 257 may alternatively be any appropriate
multichannel liquid handling head configured to deliver fluids
and/or gases. Preferably, the multichannel liquid handling head 257
comprises at least eight independent channels 258, but may
alternatively comprise any number of channels 258 configured to
aspirate and deliver fluids. The channel-to-channel pitch is
preferably variable, and in a specific example ranges between 9 mm
and 36 mm; however, the channel-to-channel pitch may alternatively
be fixed, as shown in FIG. 15. The multichannel liquid handling
head 257 also preferably provides independent z-axis control (in
the orientation shown in FIG. 14B), such that, in combination with
the gantry 256. The multichannel liquid handling head 257 is
preferably configured to couple to both large (e.g., 1 mL) and
small (e.g., between 100 and 300 .mu.L) pipette tips, and in a
specific example, has a precision of at least 6% using small
disposable pipette tips and a precision of at least 2% using large
disposable pipette tips when dispensing essentially the entire tip
volume. Alternatively, the multichannel liquid handling head 257
may be configured to couple to any object configured to facilitate
aspiration and delivery of fluids. Preferably, the multichannel
liquid handling head 257 provides independent control of the
channels 258, with regard to volumes of fluid aspirated or
delivered, fluid dispensing rates, and/or engaging and disengaging
pipette tips. Alternatively, the multichannel liquid handling head
257 may not provide independent control of the channels 258, such
that all channels 258 of the multichannel liquid handling head 257
are configured to perform identical functions simultaneously.
Preferably, the multichannel liquid handling head 257 is configured
to aspirate and deliver both liquids and gases, but alternatively,
the multichannel liquid handling head 257 may be configured to only
aspirate and deliver liquids. Preferably, the multichannel liquid
handling head 257 provides at least one of liquid level detection,
clot detection, and pipette tip engaging/disengaging detection for
each of the channels 258; however, the multichannel liquid handling
head 257 may alternatively not provide liquid level detection, clot
detection, and pipette tip engaging/disengaging detection for each
of the channels 258.
[0103] In one embodiment, the multichannel liquid handling head 257
is configured to couple to at least one filter 260, which functions
to pre-filter liquids being aspirated and/or dispensed by the
liquid handling arm 255, and is preferably a custom filter 260
configured to couple to a pipette tip, but may alternatively be any
appropriate filter configured to couple to the liquid handling arm
255 and filter liquids being aspirated and/or dispensed by the
liquid handling arm 255.
[0104] An embodiment of a custom filter 260, as shown in FIG. 22,
comprises a first end 261 configured to couple to a pipette tip, a
pointed second end 262, a void 263 coupled to the first end 261 and
the pointed second end 262, and a filter membrane 264 subdividing
the void 263. The first end 261, as shown in FIG. 22, preferably
comprises a tapered channel configured to provide a friction fit
with a pipette tip; however, the first end may alternatively not
comprise a tapered channel and may be configured to couple to a
pipette tip using any appropriate means. The pointed second end 262
is preferably sharp and configured to pierce an object, such as a
foil seal; additionally, the pointed second end 262 is preferably
at least as long as required to dispense into a well 113 of the
capture plate 110. The void 263 preferably defines a conical region
defined by the filter membrane 264, wherein the conical region is
configured to divert a fluid within the filter 260 toward the
pointed second end 262; however, the void 263 may not include a
conical region. The filter membrane 264 functions to filter a fluid
aspirated by the multichannel liquid handling head 257, and is
configured to subdivide the void 263 to define a conical region;
however, the filter membrane 264 may alternatively not define a
conical region of the void 263. In one embodiment, in the
orientation shown in FIG. 22, the region of the void 263 below the
filter membrane 264 may have a volumetric capacity of between 200
ul and 1 mL; however, the region of the void 263 below the filter
membrane may alternatively have any appropriate volumetric
capacity.
[0105] A set of filters 260 may further be configured to be
received and delivered by a filter holder 269, as, shown in FIG.
23. A specific embodiment of a filter holder 269 comprises a set of
24 tapered holes with an 18 mm center-to-center pitch, arranged in
six rows of four holes. The specific embodiment of the filter
holder 269 is also compliant with ANSI and SLAS standards, has
dimensions of 127.75.times.85.5.times.14.35 mm, and is stackable
with other specific embodiments of the custom filter holder 269.
Alternatively, the filter holder 269 may be any appropriate filter
holder 269 configured to receive and deliver a set of filters 260,
as is readily known by those skilled in the art.
1.5.1 Fluid Handling System--Syringe Pump
[0106] The syringe pump 265 of the liquid handling system 250 is
coupled to a wash solution source 266, a release solution source
267, a source of air 268, and flexible tubing 291, and functions to
deliver a wash solution, a release solution, and air through a
valve to the molecular diagnostic module 130 to facilitate
isolation and purification of nucleic acids from a set of magnetic
bead-samples. The flexible tubing 291 is preferably coupled at a
first end to the syringe pump, and at a second end to a nozzle 149
coupled to the linear actuator 146 of the molecular diagnostic
module 130, as shown in FIG. 14C. As stated earlier, an extended
configuration 146b of the linear actuator 146 is configured to
couple the nozzle 149 to a fluid port 222 of a microfluidic
cartridge 210 within the molecular diagnostic module 130, such that
the wash solution, release solution, and air can be delivered to
the microfluidic cartridge 210 at appropriate stages. A specific
embodiment of the syringe pump 265 comprises a 4-way valve, is able
to pump 20-500 .mu.L of fluids or air through the 4-way valve at
flow rates from 50-500 .mu.L/min, can couple to syringes with
between 1 mL and 10 mL capacities, and has a precision of at least
5% with regard to fluid or air delivery. Alternatively, the syringe
pump 265 may be any appropriate syringe pump 265 or fluid delivery
apparatus configured to deliver a wash solution, a release
solution, and air to the molecular diagnostic module 130, as is
readily known by those skilled in the art.
1.6 System--Additional Elements
[0107] The system 100 may further comprise a tag reader 271, which
functions to read barcodes, QR codes and/or any other identifying
tags of the system 100. Preferably, the tag reader 271 is coupled
to the liquid handling system 250, such that the tag reader 271 is
configured to read tags on puncturable foil seals 115, 195 or tags
located on any element of the system 100 accessible by the liquid
handling system 250; however, the tag reader 271 may alternatively
not be coupled to the liquid handling system 250. In one
alternative embodiment of the system 100, the tag reader 271 may be
a standalone unit that is configured to be manipulated by a user to
scan tags or labels located on elements of the system 100.
[0108] The system 100 may also further comprise a controller 272
coupled to at least one of the capture plate module 120, the
molecular diagnostic module 130, the liquid handling system 250,
and the tag reader 271, and functions to facilitate automation of
the system 100. In a variation wherein the controller 272 is
coupled to the capture plate module 120, the controller 272
preferably functions to automate heating of a capture plate 110,
which facilitates lysing of biological samples within the capture
plate 110 and binding of nucleic acids within the capture plate 110
to magnetic beads 119 of the capture plate 110. In a variation
wherein the controller 272 is coupled to the molecular diagnostic
module 130, the controller 272 preferably functions to automate
reception of a microfluidic cartridge, heating of biological
samples within the molecular diagnostic module 130 and the
detection chambers 213, occlusion of fluidic pathways 220 by the
valve actuation subsystem 170, and analysis of a set of nucleic
acid-reagent mixtures by the optical subsystem 180. In a variation
wherein the controller 272 is coupled to the liquid handling system
250, the controller 272 preferably functions to automate
aspiration, transfer, and delivery of fluids and/or gases to
different elements of the system 100. In a variation wherein the
controller 272 is coupled to the tag reader 271, the controller
preferably functions to automate reading of tags by the tag reader
271, and may further function to facilitate transfer of information
from the tags to a processor 273. Other variations of a controller
may function automate handling, transfer, and/or storage of other
elements of the system 100, such as capture plates 110, assay
strips 190, assay strip holders 230, assay strip carriers 240,
filters 200, filter holders 205, and/or microfluidic cartridges
210, using a robotic arm or gantry similar to that used in the
liquid handling system 250. Alternative combinations of the above
variations may involve a single controller 272, or multiple
controllers configured to perform all or a subset of the functions
described above.
[0109] The system 100 may also further comprise a processor 273,
which functions to receive and process information from a tag
reader 271, and also to receive and process data received from the
optical subsystem 180 of the molecular diagnostic module 130.
Preferably, the processor 273 is coupled to a user interface 274,
which functions to display processed and/or unprocessed data
produced by the system 100, settings of the system 100, information
obtained from a tag reader 271, or any other appropriate
information. Alternatively, the processor 273 is not coupled to a
user interface 274, but comprises a connection 275 configured to
facilitate transfer of processed and/or unprocessed data produced
by the system 100, settings of the system 100, information obtained
from a tag reader 271, or any other appropriate information to a
device external to the system 100.
[0110] As a person skilled in the art will recognize from the
previous detailed description and from the figures and claims,
modifications and changes can be made the described embodiments of
the system 100 without departing from the scope of the system
100.
2. Method for Processing and Detecting Nucleic Acids
[0111] An embodiment of a method 400 for processing and detecting
nucleic acids from a set of biological samples comprises: combining
each biological sample of the set of biological samples with a
quantity of magnetic beads to produce a set of magnetic bead-sample
mixtures S410; heating the set of magnetic bead-sample mixtures to
produce a set of nucleic acid-magnetic bead samples S420;
transferring each nucleic acid-magnetic bead sample of the set of
nucleic acid-magnetic bead samples to a corresponding fluidic
pathway of a set of fluidic pathways S430; producing a set of
nucleic acid volumes from the set of nucleic acid-magnetic bead
samples S440; combining each nucleic acid volume of the set of
nucleic acid volumes with a molecular diagnostic reagent of a set
of molecular diagnostic reagents to produce a set of nucleic
acid-reagent mixtures S450; transferring each of the set of nucleic
acid-reagent mixtures, through the corresponding fluidic pathway of
the set of fluidic pathways, to a detection chamber of a set of
detection chambers S460; and receiving light from the set of
nucleic acid-reagent mixtures S470. The method 400 may further
comprise generating a set of data based on light received form the
set of nucleic acid-reagent mixtures S480. The method 400 functions
to isolate and extract a set of nucleic acid volumes from a
biological sample, and to facilitate analysis of the nucleic acid
volumes according to at least one molecular diagnostic
protocol.
[0112] Step S410 recites combining each biological sample of the
set of biological samples with a quantity of magnetic beads to
produce a set of magnetic bead-sample mixtures, and functions to
prepare a set of biological samples to be lysed and combined with
magnetic beads. For each biological sample, Step S410 preferably
comprises aspirating a portion of the volume of the biological
sample from a sample container (possibly containing an aqueous
solution prior to addition of biological sample), and transferring
the portion of the biological sample to a well containing a set of
magnetic beads. Alternatively, for each biological sample, Step
S410 may comprise aspirating the entire volume of the biological
sample from a sample container, and transferring the volume of the
biological sample to be combined with a set of magnetic beads.
Preferably, all biological samples in the set of biological samples
are aspirated and combined with the magnetic beads in the wells
simultaneously using a multichannel fluid delivery system; however,
all biological samples in the set of biological samples may
alternatively be aspirated and combined with a set of magnetic
beads non-simultaneously. The magnetic beads are preferably polymer
beads, precoupled with a ligand for binding to a nucleic acid, and
comprising a superparagmagnetic component. Additionally, the
magnetic beads may be treated to be positively charged. However,
the magnetic beads may alternatively be any appropriate magnetic
beads configured to facilitate biomagnetic separation.
[0113] In addition to combination with magnetic beads, Step 410 may
further include combining each biological sample of the set of
biological samples with a lysing enzyme (e.g. proteinase K), and a
sample process control comprising two or more nucleic acid
sequences (i.e., one for DNA and one for RNA) to be included with
each sample. This allows biological samples to effectively lysed,
which releases waste components into a wash solution, and allows
nucleic acids to bind to magnetic beads. This additionally allows
the sample process control to be later detected, as a check to
verify the accuracy of a molecular diagnostic assay being
performed.
[0114] In a first variation of Step S410 for one biological sample,
as shown in FIG. 16A, a volume of the biological sample is
aspirated and combined with a set of magnetic beads. In the first
variation of Step S410, a set of different biological samples may
thus be aspirated simultaneously, and each biological sample may be
transferred to an individual well to be combined with a set of
magnetic beads to produce a set of magnetic bead-sample mixtures.
In the first variation of Step S410, all magnetic bead-sample
mixtures in the set of magnetic bead-sample mixtures are
substantially non-identical in composition. In a second variation
of Step S410, as shown in FIG. 16B, a volume of a stock biological
sample is aspirated, and portions of the volume of the stock
biological sample are transferred to multiple wells to be combined
with multiple sets of magnetic beads to produce a set of magnetic
bead-sample mixtures. In the second variation of Step S410, all
magnetic bead-sample mixtures in the set of magnetic bead-sample
mixtures are substantially identical in composition. Other
variations of Step S410 may additionally or alternatively comprise
filtering at least one biological sample of the set of biological
samples S415 prior to combining each biological sample of the set
of biological samples with a quantity of magnetic beads.
[0115] In a specific example of Step S410, a multichannel liquid
handling system aspirates approximately 1 mL of each of a set of
biological samples in aqueous buffer using a set of 1 mL pipette
tips, couples each of the pipette tips to a custom 13 mm diameter
filter, punctures a foil seal 115 of a capture plate at a set of
wells, wherein each well of the set of wells contains a set of
magnetic beads, and dispenses each aspirated volume of a biological
sample into a well of the capture plate containing a set of
magnetic beads, and disposes of the tip/filter combination. In the
specific example of Step S410, the multichannel liquid handling
system then picks up new disposable tips and aspirates and
dispenses the contents of each well of the set of wells of the
capture plate at least three times to mix the contents, and then
disposes of the set of pipette tips and filters.
[0116] Additionally or alternatively, the method 400 can include
combining each biological sample of the set of biological samples
with buffer solution S405 prior to combining each biological sample
of the set of biological samples with a quantity of magnetic beads,
which functions to further decrease sample preparation burden on an
entity processing and/or analyzing the set of biological samples.
Combining each biological sample of the set of biological samples
with buffer solution can be performed using a buffer plate
comprising a set of wells, each well containing buffer solution,
wherein the buffer plate is included in an embodiment of the system
100 described above; however, Block S405 can additionally or
alternatively be implemented using any other suitable portion of
the system 100 described above. In one variation, a liquid handling
system can be automatically configured to aspirate the set of
biological samples and mix the biological samples with the buffer
solution within the buffer plate, by dispensing the biological
samples into the buffer plate and aspirating and delivering the
samples combined with buffer one or more times. In another
variation, combination of the set of biological samples with buffer
solution can be implemented by the liquid handling system away from
the buffer plate, for instance, by aspirating the set of biological
samples, aspirating the buffer solution, and then mixing the
biological samples with the buffer solution within the liquid
handling system. In other variations, the biological samples can be
combined with buffer solution in any other suitable manner (e.g.,
altogether with mixing the biological sample with magnetic beads).
In relation to the specific example of Block S410 described above,
the multichannel liquid handling system can be configured to
aspirate each of the set of biological samples with a set of
pipette tips, dispense each of the set of biological samples into a
corresponding well of a set of wells containing buffer solution,
mix each of the set of biological samples with the buffer solution
using the set of pipette tips, while preventing cross contamination
across the set of samples by way of the set of pipette tips, couple
the set of pipette tips to a custom filter, and push the biological
samples mixed with buffer solution into wells of a capture plate,
each well of the capture plate containing a set of magnetic beads.
As such, mixing of biological samples with buffer can be performed
in an automated manner using an embodiment of the system 100
described above, prior to mixing the biological samples with
magnetic beads in Block S410.
[0117] Step S420 recites heating the set of magnetic bead-sample
mixtures to produce a set of nucleic acid-magnetic bead samples,
and functions to incubate the set of magnetic bead-sample mixtures
in order to lyse biological matter, and release nucleic acids to be
bound to magnetic beads. Preferably, Step S420 comprises heating a
capture plate containing the set of magnetic bead-sample mixtures
for a specified amount of time at a specified temperature, and may
additionally include cooling the set of magnetic bead-sample
mixtures. In a specific example, Step S420 comprises heating a
capture plate containing the set of magnetic bead-sample mixtures
using a capture plate module, wherein the capture plate module is
configured to cradle and controllably heat wells containing the set
of magnetic bead-sample mixtures. Step S420 may alternatively
comprise incubating the set of magnetic bead-sample mixtures using
any appropriate method and/or system as is known by those skilled
in the art. Finally, Step S420 may be omitted in embodiments of the
method 400 involving samples that do not require heating.
[0118] Step S430 recites transferring each nucleic acid-magnetic
bead sample of the set of nucleic acid-magnetic bead samples to a
corresponding fluidic pathway of a set of fluidic pathways, and
functions to isolate each of the set of nucleic acid-magnetic bead
samples within separate pathways for further processing.
Preferably, all nucleic acid-magnetic bead samples in the set of
nucleic acid-magnetic bead samples are transferred simultaneously
to the set of fluidic pathways, but alternatively, each nucleic
acid-magnetic bead sample in the set of magnetic bead-samples may
be transferred to a corresponding fluidic pathway independently of
the other nucleic acid-magnetic bead samples. In addition,
preferably the entire volume, or substantially all of the volume,
of the nucleic acid-magnetic bead sample is transferred to the set
of fluidic pathways, without magnetically isolating magnetic beads
and removing supernatant fluids prior to transferring each nucleic
acid-magnetic bead sample of the set of nucleic acid--magnetic bead
samples to a corresponding fluidic pathway of a set of fluidic
pathways.
[0119] Step S430 may further comprise occluding at least one
fluidic pathway of the set of fluidic pathways at a subset of a set
of occlusion positions S432, which functions to define at least one
truncated fluidic pathway. Preferably, Step S432 comprises defining
at least one truncated fluidic pathway passing through at least one
of a heating region and a magnetic field; however, Step S432 may
alternatively not comprise defining a truncated fluidic pathway
passing through at least one of a heating region and a magnetic
field.
[0120] In a specific example of Step S430, the multichannel liquid
handling subsystem of Step S410 transfers a set of nucleic
acid-magnetic bead samples to a set of fluidic pathways of a
microfluidic cartridge aligned within a molecular diagnostic
module, wherein the microfluidic cartridge comprises an elastomeric
layer in contact with the set of fluidic pathways. Manipulation of
the elastomeric layer at a subset of a set of occlusion positions
by a valve actuation subsystem of the molecular diagnostic module
defines a set of truncated fluidic pathways crossing a heating
region and a magnetic field, such that each nucleic acid-magnetic
bead sample in the set of nucleic acid-magnetic bead samples is
isolated within a truncated fluidic pathway of the set of truncated
fluidic pathways.
[0121] Step S440 recites producing a set of nucleic acid volumes
from the set of nucleic acid-magnetic bead samples, and functions
to separate nucleic acid volumes from the set of nucleic
acid-magnetic bead samples. Step S440 preferably reduces a
concentration of unwanted matter from the set of biological samples
being processed, to an acceptable level; however, Step S440 may
alternatively entirely remove substantially all unwanted substances
from the set of biological samples being processed. Step S440
preferably includes providing a magnetic field S441, such that each
fluidic pathway in the set of fluidic pathways is configured to
cross the magnetic field. Preferably, the set of nucleic
acid-magnetic bead samples is captured and isolated within portions
of the set of fluidic pathways crossing the magnetic field. Step
S440 may further comprise providing a heater configured to span a
heating region of the set of fluidic pathways S442, but may
alternatively comprise providing multiple heaters or altogether
omit providing a heater. In embodiments wherein multiple heaters
are provided, each heater is preferably independent to allow
independent control of heating time and temperature for each
sample. Step S442 functions to provide a heater, which, in
combination with a release solution that provides a pH shift,
facilitate a rapid and efficient unbinding of the nucleic acids
from magnetic beads.
[0122] Step S440 may further comprise occluding at least one
fluidic pathway of the set of fluidic pathways at a subset of a set
of occlusion positions S443 (and opening a previously occluded
channel), which functions to define at least one truncated fluidic
pathway containing a nucleic acid-magnet bead sample and coupled to
a source for delivery of a wash solution and a release solution.
Preferably, Step S443 comprises defining at least one truncated
fluidic pathway coupled to a waste chamber and to a fluid port,
which functions to facilitate washing of at least one nucleic
acid-magnetic bead sample in the set of nucleic acid-magnetic bead
samples, and releasing of at least one nucleic acid volume from the
set of nucleic acid-magnetic bead samples. Step S440 may
additionally comprise delivering a wash solution through a portion
of at least one fluidic pathway S444, such as the truncated fluidic
pathway defined in Step S443, and delivering a release solution
through a portion of at least one fluidic pathway S445, such as the
truncated fluidic pathway defined in Step S443. Step S444 functions
to wash at least one nucleic acid-magnetic bead sample in the set
of nucleic acid-magnetic bead samples, and Step S445 functions to
release at least one nucleic acid volume from the set of nucleic
acid-magnetic bead samples. The heater provided in Step S442 may be
activated after Step S445 to induce a pH shift.
[0123] In a specific example of Step S440, the set of fluidic
pathways containing a set of nucleic acid-magnetic bead samples,
from the specific example of Step S430, is occluded at a subset of
the set of occlusion positions by a valve actuation subsystem of
the molecular diagnostic module, to define a set of truncated
fluidic pathways coupled to a waste chamber and to a shared fluid
port of the microfluidic cartridge for delivery of a wash solution
and a release solution. The liquid handling system delivers a wash
fluid through the shared fluid port to wash the set of nucleic
acid-magnetic bead samples, captured within the magnetic field, and
then delivers a release fluid through the shared fluid port to
release a set of nucleic acid volumes from the set of nucleic
acid-magnetic bead samples. In the specific example, each fluidic
pathway is washed sequentially, and the release solution is
delivered to each fluidic pathway sequentially to ensure that each
lane is provided with substantially equal amounts of wash and
release solutions. All waste fluid produced in the specific example
of Step S440 pass into the waste chamber coupled to the set of
truncated fluidic pathways.
[0124] Step S450 recites combining each nucleic acid volume of the
set of nucleic acid volumes with a molecular diagnostic reagent of
a set of molecular diagnostic reagents to produce a set of nucleic
acid-reagent mixtures, which functions to prepare the set of
nucleic acid volumes to be detected. For each nucleic acid volume
in the set of nucleic acid volumes, Step S450 preferably comprises
aspirating an entire volume of the nucleic acid volume from its
corresponding fluidic pathway, and transferring the nucleic acid
volume to a well containing a molecular diagnostic reagent.
Preferably, all nucleic acid volumes in the set of nucleic acid
volumes are aspirated and combined with molecular diagnostic
reagents simultaneously using a multichannel fluid delivery system;
however, each nucleic acid volume in the set of nucleic acid
volumes may alternatively be aspirated and combined with molecular
diagnostic reagents independently of the other nucleic acid
volumes. The molecular diagnostic reagents preferably comprise
reagents configured to analyze the set of nucleic acid volumes for
markers of at least one of gonorrhea (GC), Chlamydia (CT), herpes
simplex virus (HSV), human immunodeficiency virus (HIV), human
respiratory diseases, vaginal diseases, hepatitis C virus (HCV),
hepatitis B virus (HBV), trichonomas, group B streptococcus (GBS),
factor 2 (FII) gene, and factor five (FV) gene, but may
alternatively comprise reagents used to detect any specific nucleic
acid sequence.
[0125] In a first variation of Step S450 as shown in FIG. 16A, a
nucleic acid volume is aspirated and combined with a molecular
diagnostic reagent for a single assay. In the first variation of
Step S450, a set of nucleic acid volumes may thus be aspirated
simultaneously, and each nucleic acid volume may be transferred to
an individual well to be combined with a molecular diagnostic
reagent of a set of molecular diagnostic reagents to produce a set
of nucleic acid-reagent mixtures. In the first variation of Step
S450, all nucleic acid-reagent mixtures in the set of nucleic
acid-reagent mixtures may or may not be substantially identical in
composition, depending on the homogeneity of the biological samples
used in Step S410; however, the first variation of S450 preferably
comprises using identical molecular diagnostic reagents, such that
identical molecular diagnostic protocols analyzing identical
markers may be performed. Thus, the first variation of Step S450
encompasses running multiple identical tests from a stock
biological sample (e.g., a multiplex assay), and running identical
tests using a set of substantially different biological samples
(e.g., from different sources).
[0126] In a second variation of Step S450, as shown in FIG. 16B,
the set of nucleic acid volumes is aspirated, and each nucleic acid
volume in the set of nucleic acid volumes is combined with a
molecular diagnostic reagent of a set of molecular diagnostic
reagents. In the second variation of Step S450, the set of
molecular diagnostic reagents preferably comprises different
molecular diagnostic reagents, such that different molecular
diagnostic protocols analyzing different markers may be performed.
Thus, the second variation encompasses running multiple
substantially different tests using a stock biological sample, and
running substantially different tests using substantially different
biological samples (e.g., from different sources).
[0127] In a specific example of Step S450, a multichannel liquid
handling system aspirates approximately 18 .mu.L of each of a set
of nucleic acid volumes from the microfluidic cartridge used in the
specific example of Step S440 using a set of pipette tips,
punctures at least one foil seal 195 of at least one assay strip,
wherein each well of the at least one assay strip contains
molecular diagnostic reagents, and dispenses each aspirated nucleic
acid volume into a well of the assay strip. In the specific example
of S450, the multichannel liquid handling system then aspirates and
dispenses the contents of each well approximately 10 times to
reconstitute molecular diagnostic reagents and mix the contents of
each well.
[0128] Step S460 recites transferring each of the set of nucleic
acid-reagent mixtures, through the corresponding fluidic pathway of
the set of fluidic pathways, to a detection chamber of a set of
detection chambers, which functions to deliver the set of nucleic
acid-reagent mixtures to an isolated detection chamber for further
processing and analysis. Preferably, all nucleic acid-reagent
mixtures in the set of nucleic acid-reagent mixtures are
transferred simultaneously to the set of fluidic pathways, but
alternatively, each nucleic acid-reagent mixture in the set of
nucleic acid reagent mixtures may be transferred to a corresponding
fluidic pathway independently of the other nucleic acid reagent
mixtures. Step S460 may further comprise occluding at least one
fluidic pathway of the set of fluidic pathways at a subset of a set
of occlusion positions S462, which functions to define at least one
truncated fluidic pathway coupled to a detection chamber of a set
of detection chambers. Preferably, Step S462 comprises occluding
each fluidic pathway of the set of fluidic pathways at a subset of
a set of occlusion positions, thus defining a set of truncated
fluidic pathways, each coupled to a detection chamber.
[0129] In a specific example of Step S460, the multichannel liquid
handling subsystem of the specific example of Step S450 transfers a
set of nucleic acid-reagent mixtures, each having a volume of
approximately 16 .mu.L, back to the set of fluidic pathways of the
microfluidic cartridge of the specific example of Step S450. Each
nucleic acid-reagent mixture in the set of nucleic acid-reagent
mixtures is transferred at a rate of 50 .mu.L/minute. Manipulation
of the elastomeric layer at a subset of a set of occlusion
positions by the valve actuation subsystem of the molecular
diagnostic module defines a set of truncated fluidic pathways, each
coupled to a detection chamber, such that each nucleic
acid-magnetic bead sample in the set of nucleic acid-magnetic bead
samples is isolated within a truncated fluidic pathway of the set
of truncated fluidic pathways. In the specific embodiment the
occlusion position immediately upstream of the detection chamber
and the occlusion position immediately downstream of the detection
chamber are normally closed positions. During delivery, the
multichannel liquid handling subsystem generates pressure to cause
the elastomeric layer at the normally closed positions to deform
and allow fluid to flow through the normally closed positions. Once
the pressure drops after the detection chamber is filled and the
multichannel liquid handing subsystem ceases delivery, the
elastomeric layer is configured to overcome the pressure in the
channel and recloses, thereby sealing the normally closed
positions. The normally closed positions are then compressed using
the valve actuation subsystem during thermocycling to prevent
pressures generated during a molecular diagnostic assay to cause
the normally closed positions to leak. After the molecular
diagnostic assay is complete and the occlusion "pins" withdrawn,
the normally closed positions allow the samples and amplicons to be
trapped within detection chambers, substantially reducing the risk
of contamination of the lab or other samples.
[0130] Step S470 recites receiving light from the set of nucleic
acid-reagent mixtures, and functions to produce emission responses
from the set of nucleic acid-reagent mixtures in response to
transmission of excitation wavelength light or chemiluminescent
effects. Preferably, Step S470 comprises the ability to transmit
light including a wide range of wavelengths through a set of
excitation filters and through a set of apertures configured to
individually transmit light having single or multiple excitation
wavelengths onto the set of nucleic acid-reagent mixtures, and
receiving light through a set of emission filters, from the set of
nucleic acid-reagent mixtures. Step S470 may additionally comprise
reflecting light from the set of excitation filters off of a set of
dichroic mirrors, and transmitting light through the set of
dichroic mirrors to a set of photodetectors. A specific example of
Step S470 comprises using the optical subsystem 180 of the system
100 described above to transmit and receive light; however,
alternative variations of Step S470 may use any appropriate optical
system configured to transmit light at excitation wavelengths
toward the set of nucleic acid-reagent mixtures, and to receive
light at emission wavelengths from the set of nucleic acid-reagent
mixtures.
[0131] Step S480 recites generating a set of data based on light
received from the set of nucleic acid-reagent mixtures, which
functions to produce quantitative and/or qualitative data from the
set of nucleic acid-reagent mixtures. Step S480 may further
function to enable detection of a specific nucleic acid sequence
from the nucleic acid-reagent mixture, in order to identify a
specific nucleic acid sequence, gene, or organism. Preferably, Step
S480 includes converting electrical signals, produced by a set of
photodetectors upon receiving light from the set of nucleic
acid-reagent mixtures, into a quantifiable metric; however, S480
may alternatively comprise converting electromagnetic energy,
received by a set of photodetectors from the set of nucleic
acid-reagent mixtures, into a set of qualitative data. In one
variation of Step S480, the set of data may be processed by a
processor and rendered on a user interface; however, in other
variations of Step S480, the set of data may alternatively not be
rendered on a user interface.
[0132] The method 400 may further comprise re-running a biological
sample S490 if processing and/or analysis of the biological sample
results in less than ideal results. Preferably, Step S490 occurs if
an analysis of a biological sample is indeterminate due to machine
or user error. Additionally, Step S490 preferably occurs
automatically upon detection of a less than ideal result, but may
alternatively occur in response to a user prompt. Block S490 is
enabled due to rapid processing enabled by an embodiment of the
system 100 described above, wherein rerunning a sample without
sample degradation is feasible during the time it takes to run a
biological sample and determine if an analysis of the biological
sample has produced an indeterminate result.
[0133] Embodiments of the method 400 and variations thereof can be
embodied and/or implemented at least in part by a machine
configured to receive a computer-readable medium storing
computer-readable instructions. The instructions are preferably
executed by computer-executable components preferably integrated
with the system 100 and one or more portions of the processor 273
and/or the controller 272. The computer-readable medium can be
stored on any suitable computer-readable media such as RAMs, ROMs,
flash memory, EEPROMs, optical devices (CD or DVD), hard drives,
floppy drives, or any suitable device. The computer-executable
component is preferably a general or application specific
processor, but any suitable dedicated hardware or hardware/firmware
combination device can alternatively or additionally execute the
instructions.
[0134] The FIGURES illustrate the architecture, functionality and
operation of possible implementations of systems, methods and
computer program products according to preferred embodiments,
example configurations, and variations thereof. In this regard,
each block in the flowchart or block diagrams may represent a
module, segment, or portion of code, which comprises one or more
executable instructions for implementing the specified logical
function(s). It should also be noted that, in some alternative
implementations, the functions noted in the block can occur out of
the order noted in the FIGURES. For example, two blocks shown in
succession may, in fact, be executed substantially concurrently, or
the blocks may sometimes be executed in the reverse order,
depending upon the functionality involved. It will also be noted
that each block of the block diagrams and/or flowchart
illustration, and combinations of blocks in the block diagrams
and/or flowchart illustration, can be implemented by special
purpose hardware-based systems that perform the specified functions
or acts, or combinations of special purpose hardware and computer
instructions.
[0135] As a person skilled in the art will recognize from the
previous detailed description and from the figures and claims,
modifications and changes can be made to the preferred embodiments
of the invention without departing from the scope of this invention
defined in the following claims.
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