U.S. patent application number 12/671257 was filed with the patent office on 2010-08-19 for sample processor.
This patent application is currently assigned to Enigma Diagnostics Limited. Invention is credited to Trevor John Beckett, Martin Alan Lee, Christopher John Silk, David James Squirrell, Michael John Withers.
Application Number | 20100210010 12/671257 |
Document ID | / |
Family ID | 38529269 |
Filed Date | 2010-08-19 |
United States Patent
Application |
20100210010 |
Kind Code |
A1 |
Lee; Martin Alan ; et
al. |
August 19, 2010 |
SAMPLE PROCESSOR
Abstract
An integral magnet and heater device comprising a heating
element with integral permanent magnet for use in a sample
preparation or analysis apparatus. Also disclosed is a sheath
adapted to cover a reusable processing component, such as a heater
or sonicator, of an apparatus during use. Other associated aspects
of the system are described and claimed.
Inventors: |
Lee; Martin Alan;
(Wiltshire, GB) ; Squirrell; David James;
(Wiltshire, GB) ; Withers; Michael John;
(Cambridge, GB) ; Beckett; Trevor John;
(Cambridge, GB) ; Silk; Christopher John;
(Cambridge, GB) |
Correspondence
Address: |
POLSINELLI SHUGHART PC
700 W. 47TH STREET, SUITE 1000
KANSAS CITY
MO
64112-1802
US
|
Assignee: |
Enigma Diagnostics Limited
Wiltshire
GB
|
Family ID: |
38529269 |
Appl. No.: |
12/671257 |
Filed: |
August 1, 2008 |
PCT Filed: |
August 1, 2008 |
PCT NO: |
PCT/GB2008/002630 |
371 Date: |
March 11, 2010 |
Current U.S.
Class: |
435/288.3 ;
219/552 |
Current CPC
Class: |
G01N 2035/00376
20130101; G01N 2035/103 20130101; B01L 3/5029 20130101; G01N
35/1079 20130101; B01L 9/00 20130101; B01L 2200/16 20130101; B01L
2200/141 20130101; B01L 2300/0867 20130101; G01N 2035/00534
20130101; B01L 2200/025 20130101; B01L 2300/0672 20130101; B01L
2300/044 20130101 |
Class at
Publication: |
435/288.3 ;
219/552 |
International
Class: |
C12M 1/34 20060101
C12M001/34; H05B 3/10 20060101 H05B003/10 |
Foreign Application Data
Date |
Code |
Application Number |
Aug 3, 2007 |
GB |
0715171.5 |
Claims
1. An integral magnet and heater device, comprising: a heating
element; and a magnet.
2. (canceled)
3. The device according to claim 1 wherein the heating element is a
resistive device.
4. The device according to claim 1 further comprising a temperature
sensor.
5. The device according to claim 1 further comprising a temperature
control by which a temperature of the device can be set.
6. The device according to claim 1 wherein the device is an
elongate body and wherein the magnet is located on one end of the
elongate body.
7. The device according to claim 1 wherein the magnet is located
around a periphery of the heating element.
8. The device according to claim 1 wherein the magnet is a
permanent magnet, and the permanent magnet has a field strength
which remains effective up to at least 100.degree. Celsius.
9. An apparatus for sample preparation or analysis, the apparatus
comprising: one or more reagent or sample chambers; an integral
magnet and heater device comprising a heating element and a magnet,
wherein the device is movable relative to the one or more reagent
or sample chambers.
10. An apparatus for sample preparation or analysis, the apparatus
comprising: one or more reagent or sample chambers; a reusable
processing component; and a sheath adapted to cover the reusable
processing component during use.
11. The apparatus according to claim 10 wherein the sheath is
provided as a moveable component on the apparatus.
12. The apparatus according to claim 10 wherein the sheath is of a
thermally conducting plastic or an elastomeric material.
13. The apparatus according to claim 10 wherein the reusable
processing component comprises a heater or sonicator.
14. A sheath adapted to cover a reusable processing component of an
apparatus during use.
15. The sheath according to claim 14 wherein the processing
component is a heater or sonicator.
16. The sheath according to claim 14 wherein the processing
component is a heater and the sheath is of a thermally conducting
plastic or an elastomeric material.
Description
[0001] The present invention relates to a device for processing
samples, in particular clinical samples in preparation for
analysis, for instance by means of a nucleic acid amplification
method, in particular the polymerase chain reaction (PCR), as well
as novel elements and procedures which may be utilised in such
devices.
[0002] The analysis of fluid samples, for example clinical or
environmental samples, may be conducted for several reasons. One
current area of interest is the development of a method for
positively identifying biological material in a fluid sample, for
example a clinical or environmental sample. Such a method would
allow for early diagnosis of disease states, which in turn would
enable rapid treatment and infection control, or the identification
of environmental contaminants and the like. There are many
techniques by which very sensitive analysis of samples can be
carried out including for example nucleic acid amplification
techniques such as the polymerase chain reaction (PCR).
[0003] International patent application WO 2005/019836 discloses an
apparatus for processing a fluid sample which comprises a rotatable
platform and a moveable arm which can be raised and lowered. Te
platform is provided with chambers for housing the sample and
reagents. Components such as a cutter, heater, optical detector,
sonicator, magnet and sheath are housed in the platform and may be
releasably attached to the arm for movement relative to the
chambers. The components are used to move the sample_ and/or
reagent between chambers (such as magnets and sheaths), to pierce
the seal of chambers (such as cutters) and to carry out physical
processes on the content of the chambers (such as heaters and
sonicators).
[0004] Examples of sheaths for magnets useful in transferring
magnetic beads or particles and thus any reagents attached thereto
are described for example in WO2005/019836.
[0005] If desired also, processing components such as heaters,
sonicators etc. which may be required to treat reagents or reagent
mixtures to ensure that they are in a desired physical state and
any time during the procedure, may also be removeably housed on the
apparatus.
[0006] Where components such as sonicators or heaters are fitted to
the apparatus in this way, they are generally intended for repeated
use and for immersion with a sample, in particular a liquid sample
within a chamber.
[0007] This in itself, may cause some problems as the use of a
non-disposable device in such a way that it comes into direct
contact with a sample liquid brings the risk of contamination. Over
time, these components may become soiled or tarnished, thus
increasing the risk of such contamination.
[0008] A first aspect of the invention provides an integral magnet
and heater device.
[0009] This has the advantage that where the apparatus includes
both a magnet to facilitate removal of beads or particles from one
chamber to another, a heater may be incorporated into the magnet so
that the same element may fulfil both functions.
[0010] In one embodiment, the integral magnet and heater device
comprises a heating element and a magnet.
[0011] The heating element may be a resistive device, for example a
cartridge heater which typically comprises resistive wires embedded
in a thermally insulating material.
[0012] The integral magnet and heater device may further comprise a
temperature sensor. The device may further comprise a temperature
control by which the device can be set.
[0013] In a preferred embodiment, the device is an elongate body
and wherein the magnet is located on one end of the elongate body.
The magnet may be located at one end of the elongate body.
Alternatively, the magnet may be located around the periphery of
the heating element.
[0014] For example, heating elements may be incorporated into a
core of a bar magnet and this may be used both for the transport of
beads or particles, or the heating of vessel or chamber
contents.
[0015] The magnet is chosen which has a field strength which
remains effective up to at least 100.degree. C., which is
sufficient for aqueous reactions. Preferably the magnet is a
permanent magnet.
[0016] In any of the embodiments described above, the integral
magnet and heater device may be used both for the transport of
beads or particles, or the heating of vessel or chamber
contents.
[0017] In that instance, any disposable sheath utilised to cover
the magnet when it is deployed in the apparatus is suitably of a
heat conducting polymer as described above. Such combined
magnet/heaters form a further aspect of the invention and may be
used, for example, to expedite solvent evaporation from retained
magnetic particles; to heat liquids to aid dissolution of reagents;
to facilitate the desorption of compounds bound to the surface of
magnetic particles, (in particular, nucleic acids); for degassing
of liquids; or to effect or assist the lysis of cellular material,
e.g. to release nucleic acids.
[0018] A second aspect of the invention provides apparatus for
sample preparation or analysis, the apparatus comprising:
[0019] one or more reagents or sample chambers
[0020] an integral magnet and heater device which is movable
relative to the one or more reagent or sample chambers.
[0021] The apparatus may comprise a cartridge comprising a body
section adapted to hold a sealed sample vessel; and apparatus
adapted to receive said cartridge.
[0022] A third aspect of the present invention provides apparatus
for sample preparation or analysis, the apparatus comprising:
[0023] one or more reagent or sample chambers
[0024] a reusable processing component
[0025] and a sheath adapted to cover the reusable processing
component during use.
[0026] The applicants have found that the risk of contamination by
repeated use of a reusable processing component can be minimized by
placing a sheath, which is disposable, over the processing
component before use. In particular, where a processing component
such as a heater is used repeatedly during a particular chemical,
biochemical procedure or assay, a new disposable sheath is suitably
applied on each occasion.
[0027] The apparatus may comprise a cartridge comprising a body
section adapted to hold a sealed sample vessel; and apparatus
adapted to receive said cartridge.
[0028] Sheaths of this type are suitably removeably housed on the
cartridge housing the one or more reagent or sample chambers in a
similar manner to the mechanical elements described above, so that
they are readily available and can be accessed and positioned using
the arm in a similar fashion. Thus, for example, the sheath may be
provided with a lip or flange, able to interact with a fork on the
moveable arm of the apparatus and lifted into position around the
processing component as necessary.
[0029] After use, they may be returned to the cartridge for
disposal or disposed of directly.
[0030] The sheath is suitably made of a plastics or elastomeric
material, and where they are intended for use in conjunction with
reusable heaters, they are preferably made of a thermally
conducting plastic, for example plastic filled with boron nitride
or a commercially available "cool polymer" material, so as to
minimize any loses in heater efficiency.
[0031] A fourth aspect of the present invention provides a sheath
adapted to cover a reusable processing component of an apparatus
during use. Such sheaths form a further aspect of the
invention.
[0032] Where heaters are provided in the apparatus, they suitably
incorporate temperature sensors, so that the temperature of a
reagent or sample in which they are immersed can be determined.
When a sheath as described above is used, some calibration of the
apparatus will be necessary to ensure that these readings are
accurate.
[0033] A suitable calibration method for calibrating the heater
when used in combination with a sheath comprises the steps of
immersing the sheath containing the heater in a liquid bath (e.g.
water bath; varying the temperature of the heater; record the
temperature readings of heat sensor of the heater and the
temperature of the liquid bath; and using the correlation between
the temperature readings of the heat sensor of the heater and the
temperature of the liquid bath to provide calibration data.
[0034] The data can be used to create a calibration file that is
either used in software as a look-up table to convert the sensor
signals to accurate temperatures or to work out a mathematical
transform for the signals from the heat sensor.
[0035] In a particular embodiment, where the apparatus includes
both a magnet to facilitate removal of beads or particles from one
chamber to another, a heater may be incorporated into the magnet so
that the same element may fulfil both functions. For example,
heating elements may be incorporated into a core of a bar magnet
and this may be used both for the transport of beads or particles,
or the heating of vessel or chamber contents.
[0036] In that instance, any disposable sheath utilised to cover
the magnet when it is deployed in the apparatus is suitably of a
heat conducting polymer as described above. Such combined
magnet/heaters form a further aspect of the invention.
[0037] The cartridge also suitably contains further elements which
are useful in the subsequent procedures to which the sample are
required to be subjected.
[0038] Such further elements may include one or more reagent
chambers for holding reagents or materials required to continue the
analysis of the sample. These reagents or materials, which include
wash solutions, diluents or buffers as well as reagents used in the
subsequent procedure, are suitably predispensed into the reagent
chambers.
[0039] Where this involves sample preparation, for example to
extract nucleic acid from samples suspected of containing cells,
such reagents may include lysis reagents for example chaotrophic
salts, bacteriophages, enzymes (which may lyophilized), detergents,
antibiotics the like. Where the subsequent procedure involves
sample analysis, other reagents such as dyes, antibodies, enzymes
(including for example polymerase enzymes for PCR), buffers, salts
such as magnesium salts, may be included in further reaction
chambers on the cartridge. However, the range of possible reagents
is very large and they will be selected on a "case-by-case" basis,
depending on the nature of the chemical or biochemical reaction or
analysis or assay to which the sample is subjected.
[0040] The reagents may be present in solid or liquid form. When
they are predispensed in solid form, these may be as a solid
powder, bead, capsule or pressed tablet form, or they may be
adhered to magnetic particles or beads, such as silica beads as is
well known in the art.
[0041] Reagent chambers containing predispensed reagents are
suitably sealed for example using foil seals which may be piercable
within the apparatus using cutters. These may be integral with the
apparatus, but in a particularly preferred embodiment, a cutter is
removeably housed in the cartridge, for example within an
appropriately shaped recess or aperture in the body section, so
that it is available for use in relation to the particular chemical
or biochemical reaction, analysis or assay being carried out.
[0042] Other mechanical elements in addition to the cutter, in
particular those which may be of a disposable nature, which are
useful in or otherwise facilitate further chemical or biochemical
reaction, physical processing, analysis or assay of the sample, may
also be removeably housed on the cartridge. Such elements may
include devices required to move samples, reagents or materials
from one chamber to another, such as pipettors, magnets or sheaths
therefore, as well as small devices such as filters, stoppers,
mixers, caps etc. which may require to be introduced into chambers
in the course of the chemical, biochemical or analytical procedures
or assays.
[0043] If desired also, processing components such as heaters,
sonicators etc. which may be required to treat reagents or reagent
mixtures to ensure that they are in a desired physical state and
any time during the procedure, may also be removeably housed on the
cartridge.
[0044] Mechanical elements such as cutters, pipettors, magnets or
sheaths therefore, as well as other sheaths as detailed more fully
below, and processing components as described above, will
collectively be referred to hereinafter as "moveable
components".
[0045] In such cases, the apparatus is provided with means for
accessing these moveable components and for moving them as
necessary so that they can fulfil the required function in the
chemical, biochemical, analytical or assay procedure. In
particular, the apparatus may comprise a moveable arm which is able
to interact with any moveable component on the cartridge.
[0046] The moveable arm, which is for example, a robotic arm, is
suitably provided with a'grab device, so that it can pick up any
moveable components housed on the cartridge and lift it up out of
its associate recess or aperture. Suitable grab devices are known
in the art. The may include forked elements which are arranged to
removeably interact with appropriately positioned flanges on the
moveable components, but may also comprise controllable grabbing
arms, able to close around an exposed upper portion of one of the
moveable components. Again, the moveable components may include
particular adaptations such as flanges or recesses which are
arranged to interact with grabbing arms to facilitate movement.
[0047] The apparatus is designed such that a moveable component
held on the arm may be positioned above an appropriate reaction or
reagent chamber within the cartridge. The arm may be moveable
laterally as well as vertically in order to achieve this. However,
in a particular embodiment, the arm itself is moveable only in a
single dimension which is vertically, and a transport means for the
cartridge is provided, suitably as part of the apparatus, so as to
allow it to be moved in a lateral direction that the arm may be
positioned directly above each element on the cartridge including
reaction chambers, reagent chambers as well as moveable components,
so as to allow the desired sequence of events to occur. Where the
arm may be moveable laterally as well as vertically, the
requirements for the horizontal transport means for the cartridge
may be reduced.
[0048] When electrically operated processing components such as a
heater or sonicators are supplied in this way, it may be useful if
the connection between the processing component and the arm of the
apparatus were also to provide an electrical connection sufficient
to provide power to the processing component during use. However,
such elements may be more conveniently housed within the apparatus
itself, and arranged to be delivered for example to the appropriate
chamber on the cartridge at the required time. The arm itself or an
adjunct to the arm on which the processing component such as the
heater or sonicator is fitted, may be used in order to ensure that
the component can be positioned as necessary in relation to a
chamber on the cartridge.
[0049] The apparatus may also comprise devices such as thermal
cyclers, optical readers such as fluorimeters, as well as data
processing devices arranged to collect, analyse and/or record
signals from any chamber within the cartridge or apparatus. The
selection and arrangement of suitable devices within the apparatus
will depend upon the nature of the chemical and biochemical
reaction or assay being conducted, and will be within the ambit of
the skilled person.
[0050] The inclusion of multiple moveable components and chambers
on a cartridge opens up the possibility that the sample preparation
and/or analysis may be carried out in a largely self-contained unit
comprising the cartridge. Such units, including where all chambers
are moveable components, may be readily disposable after use to
avoid further contamination risks. Furthermore, by conducting an
assay in a single cartridge, it is possible to reduce the risk of
errors in sample labelling since the cartridge itself may be
labelled at the time of introduction of the sample, for example
using a standard bar code labelling system, and the label will
remain with the sample throughout the analytical procedure.
[0051] All processes are suitably carried out automatically by
programming the apparatus to move the relevant components, reagents
etc. into contact with each other in an appropriate sequence. For
example, as described in WO2005/019836 the sample can be subject to
a nucleic acid extraction procedure, followed by a PCR reaction.
However, many other procedures in which safe sample delivery is
required may be undertaken using the invention by appropriately
designing the apparatus and programming it accordingly. The
application of such robotic techniques is well known in the
art.
[0052] When the process includes a thermal cycling step such as a
PCR, the apparatus will suitably include a thermal cycling device.
The vessel in which the thermal cycling is carried out may be
positioned on the cartridge if required. However, alternatively,
where the arrangement of the cartridge and the apparatus is such
that the preparation of the sample only is carried out on the
cartridge, a particularly suitable arrangement is that the prepared
sample ends up in a removeable reaction chamber, which is then
transferred to a specific thermal cycling area (as illustrated for
example in WO2005/019836). However, this may not always be
necessary and the incorporation of a chamber which may be thermally
cycled on the cartridge would be advantageous in that it would
allow further simplification of the apparatus.
[0053] Use of electrically conducting polymer as a heater for
thermal cycling in particular in PCR reactions, as described and
claimed in WO 9824548 (the content of which is incorporated herein
by reference), provides a particularly compact and versatile system
for use in conjunction with the system of the present application,
since the ECP may be readily incorporated into a reaction chamber
which is housed, if necessary removeably housed) on the
cartridge.
[0054] Generally, the ECP is used to coat a reaction vessel which
comprises essentially two parts, a relatively wide-mouth upper
section for receiving the sample, and a lower sealed capillary tube
which then acts as the reaction vessel. At least the lower sealed
capillary tube comprises ECP which effectively acts as a highly
controllable resistance heater, when electrical contacts are placed
across it.
[0055] The invention will now be particularly described by way of
example with reference to the accompanying diagrammatic drawings in
which:
[0056] FIG. 1 shows a plan view of a cartridge useful in the system
of the invention;
[0057] FIG. 2 is a schematic perspective view of a cartridge useful
in the system of the invention which is about to receive a sample
vessel;
[0058] FIG. 3 is a perspective view of the cartridge of FIG. 2 with
a sample vessel in place within the holder;
[0059] FIG. 4 is a perspective view of the cartridge of FIG. 3 in a
"closed" position;
[0060] FIG. 5 is an end view of the cartridge of FIG. 4;
[0061] FIG. 6 is an end view of the cartridge of FIG. 3;
[0062] FIG. 7 is a schematic diagram showing how a cartridge of the
invention may be introduced into a apparatus in which a chemical,
biochemical or other type of assay or processing may be
conducted;
[0063] FIG. 8 illustrates a cartridge of the invention in position
in a receiving section of an apparatus;
[0064] FIG. 9 is a schematic diagram illustrating a system for
filling a capillary tube, which forms a further aspect of the
invention;
[0065] FIG. 10 illustrates a sheath for use in the invention;
[0066] FIG. 11 illustrates a side view of a combined heater and
magnet;
[0067] FIG. 12 is a cross section of the lower part of the combined
heater and magnet of FIG. 11; and
[0068] FIG. 13 illustrates a side view of a heating element for use
in the combined heater and magnet of FIGS. 11 and 12.
[0069] The cartridge shown in FIG. 1 includes a body section (1)
which is of a rigid plastic material and is of generally oblong
section. A clip feature is provided to facilitate location of the
cartridge when it is placed in the instrument. A central
longitudinal channel (2) is provided in the upper surface (3) of
the body section (1). The channel (2) is open at one end but is
does not extend the full length of the body section (1) so that it
terminates in an end ridge (4) of the body section. The channel (2)
has a generally curved base (5) and is shaped so that it could
accommodate a tube (6) with a sealing cap (7). The channel 2 is
inclined downwards towards the ridge (4) so that a liquid sample
contained within the tube (6) will flow towards the cap (7).
[0070] In the illustrated embodiment, the tube accommodates a swab
(8) which is fixed to the cap (7) by way of a support (9).
[0071] The cartridge also contains a reaction chamber (10). A
piercing needle (not shown) extends between the chamber (10)
towards the cap (7) with a piercing tip at the end adjacent the cap
(7). The cap (7) suitably includes a piercable membrane (11) (FIG.
2) in the upper surface thereof.
[0072] When the tube (6) is in position in the channel (2), the cap
(7) is sufficiently far removed from the piercing needle to ensure
that it is not breached. However, the cartridge (1) is designed to
be positioned within an apparatus (not shown), which is provided
with an actuator able to apply pressure to the base of the tube (6)
in the direction of the arrow. This forces the membrane (11) of the
cap (7) against the piercing needle, which passes through the
membrane (11) and thus breaches the seal.
[0073] Any liquid within the tube (6) is able to flow out through a
channel in the needle into the reaction chamber 10, where it may be
subject to further processing. However, no operator contact with
the contents of the tube (6) has taken place at this point and so
the risk of contamination is minimised.
[0074] The cartridge (1) also includes in side sections a number of
components or elements which may be utilised in an automated
analytical process. For instance, it contains a number of foil
sealed reservoirs (12) which may contain liquid reagents such as
buffers, washes etc. which may be required for the desired
processing of a sample. Others (13) may contain reagents such as
solid reagents such as FOR beads useful in the subsequent
processing of the sample.
[0075] In addition in this instance, the cartridge includes a
series of movable components including two pipettors 14, a stopper
15 and a sheath 16 which may fit for example over a magnet used to
move magnetic reagent beads from one chamber to another on the
cartridge as required. These moveable components are accommodated
within appropriately shaped apertures in the upper surface (3) of
the body section (1). They are arranged so that an upper region
projects above the upper surface (3) so that they are accessible
for a grabbing arm of an apparatus. They may be provided with
suitable annular flanges to facilitate this, or to assist in the
lifting operation, for example as described in WO2005/019836.
[0076] In this case also, there is a provided in the cartridge (1),
a reaction vessel (17) which is coated with an electrically
conducting polymer, and so which, when connected to a suitable
electrical supply, can subject the contents to a thermal cycling
procedure such as that required for PCR.
[0077] The arrangement of this vessel will be discussed in more
detail hereinafter in relation to other illustrated
embodiments.
[0078] The cartridge illustrated in FIG. 2 contains many common
elements although these are slightly differently arranged to suit
the particular apparatus and chemical, biochemical or analytic
procedure or assay being carried out. However, in this case, a
holder (18) for the tube (6) is provided. The holder (18) is also
tubular in shape and is capable of holding the tube (6) such that
the cap (7) abuts against the end (FIG. 3).
[0079] The holder (18) may be retained against the cartridge body
(1) in an upright sample vessel receiving position by means of a
clasp (20) (FIGS. 5 and 6) disposed at the free end of the channel
(2).
[0080] Once a tube (6) has been loaded into the holder (18), it is
removed from the clasp (20), and inserted into the channel (2). A
flange (19) provided on the side of the holder (18) is arranged to
engage in a snap fit locking arrangement with a corresponding
groove in the ridge (4) of the body section (1), but only if the
tube (6) is snugly fitted into the holder (18) (FIG. 4).
[0081] At this point, the tube (6) and the cap (7) are
substantially completely encased within the cartridge and holder
and so are not accessible for fracture etc. A space (22) for a
label for a bar-code reader to identify the cartridge and a window
(23) to allow a bar-code on the sample tube to be read may be
provided on the flange (19) and holder (18) respectively.
[0082] Sample labels may be applied at this point to the cartridge
for example bar code labels which may be applied to an end region
(21) of the body section (1), so as to facilitate tracking of the
sample through the analytical procedure.
[0083] The base of the holder (18) includes a small aperture (24)
(FIG. 6). The aperture (24) is shaped to allow an actuator of the
apparatus into which the cartridge is introduced to pass through
and so urge the tube (6) towards the piercing needle provided at
the region of the ridge (4)
[0084] Once the actuator has passed through the aperture (24), the
cartridge is effectively "locked" and cannot then be opened. The
actuator is then withdrawn whilst the sample tube remains in
position at least until the end of the analytical procedure.
[0085] The cartridge (1) is shaped so that it may be received into
a receiving section of a suitable apparatus. This is illustrated
schematically in FIG. 7. In that case, the cartridge receiving
section of the apparatus (36) comprises a support (37) provided
with a recess (38), into which the cartridge (1) snugly fits. The
support (37) is retractable into the body of the apparatus (36),
for processing. The support (36) is itself moveable (see arrows) so
as to align any particular part of the cartridge (1) with an
interacting element (39), which may be moveable in a vertical
direction.
[0086] A similar arrangement is illustrated in FIG. 8. In this
case, the cartridge (1) is provided with a lip (40) which engages
the upper surface of the support (37) when the cartridge is in
position within the recess (38). The holder (18) is arranged so
that when the support (37) is retracted into the body of the
apparatus, the actuator for opening the tube (6) and can enter
through the aperture (11) to release sample into the sample vessel
(10) prior to the processing procedure. If required, locking or
other engagement means may be provided to fix the cartridge (1) in
position on the support (37).
[0087] Thus in use, a sample is collected for example for chemical,
biochemical analysis, investigation or assay. If the sample is a
liquid sample, it is suitably placed directly in a tube (6) which
is sealed with a cap (7). Preferably the volume of the sample is
known or is measured, in particular if the nature of the
investigation being carried out is qualitative in nature. The
sample tube may be inscribed with maximum and minimum fill lines to
facilitate the dispensing of the liquid sample and to provide a
means of checking that the sample volume is within the required
limits. If the sample has been collected on a swab, then the swab
(8) itself is placed in the tube together with a suitable and
preferably known volume of eluent and the tube (6) is then sealed
with a cap (7). The tube is then suitably shaken to ensure that any
sample is transferred from the swab (8) to the eluent, although
this may not be necessary if the volume of the liquid is sufficient
to ensure that the swab remains immersed in the liquid.
[0088] Then either directly, or when it reaches a laboratory, the
tube (6) is placed in a holder (18) of a cartridge. The holder is
then inserted into the channel (2) of the body section (1) of a
cartridge and the cartridge itself is labelled, before being placed
into an appropriate cartridge receiving section of an apparatus
(designed to effect the necessary procedures so as to effect the
chemical, biochemical or analytical procedures or assays on the
sample.
[0089] At this point, an actuator on the apparatus is caused to
pass through the aperture (24) in the base of the holder (18) so as
to urge the tube (7) towards the hollow piercing needle at the
ridge end of the cartridge. Sufficient pressure is applied to the
tube (6) by the actuator (24) to ensure that the rubber seal (11)
in the cap (7) is breached by the needle.
[0090] Because the tube (6) is inclined downwards towards the ridge
(4), the liquid contained therein will run through the hollow
piercing needle directly into the reaction chamber (10) on the
cartridge.
[0091] The apparatus is then able to effect processing, for example
using robotic procedures known in the art. A vertically moveable
arm is suitably used to effect the processing, whilst the cartridge
is moveable, for example by Cartesian motion, so that the
appropriate chamber or component on the cartridge is aligned with
the arm at any one time.
[0092] The possibility for assay design using this procedure is
limitless, as all that it is necessary to do in any particular case
is to ensure that reagent containers on the cartridge and that
suitable other components such as the moveable components described
hereinbefore, are provided either on the cartridge or integrated
appropriately into the apparatus.
[0093] A particular example of such a procedure is illustrated in
WO2005/019836.
[0094] To summarise that procedure however, a sample within the
chamber 10 which is known or suspected of containing cells of
interest is subject to cell lysis. This may be achieved for example
by preloading the chamber 10 with a chemical lysis agent such as
guanidine hydrochloride, by adding such a reagent taken from a
reagent container for example using a pipettor 14, by introduction
of a sonicator which is suitably integral with the apparatus or a
combination of these. Where reagents are obtained from a sealed
container 12 on the cartridge, they may be accessed following
piercing of the foil lids with a cutter, which itself may be a
moveable component on the cartridge or an integral part of the
apparatus.
[0095] Magnetic beads which are suitably coated with a binding
agent such an antibody specific for a particular target analyte or
nucleic acid generically, such as "Magnesil.RTM." silica beads are
then introduced, for example using a magnet which is inserted into
a sheath 16 and brought into contact with beads when it attraction
is required (for example to pick the beads out of a container) and
removed from the sheath when the beads are required to be
deposited, for instance once the sheath has been positioned inside
the reaction chamber 10.
[0096] After allowing the analyte such as any nucleic acid to
become adhered to the beads, they may be removed from the reaction
chamber (10) and placed into a different reaction chamber, which
may have been foil sealed until the seal was broken by a suitable
cutter before addition of the analyte. The beads may be moved
through one or more wash chambers, optionally present on the
cartridge, at this time if required.
[0097] Analyte may then be eluted from the beads for example by
adding the beads to an eluent, which is preferably hot, contained
in a chamber (12). Heating of the eluent may take place by
introducing a heater provided on the apparatus, which is preferably
encased within a protective disposable sheath 16 as described
above. However, in the event that it is not, it may be subject to
washing steps using wash liquids which may be contained in reagent
chambers which are optionally on the cartridge.
[0098] A sheath (16) is shown in more detail in FIG. 10 and
comprises a hollow cylindrical body (40), sealed at its lower end
(42). The upper end (44) has an opening, surrounded by a flange
(46) which is able to interact with a fork on the moveable arm of
the apparatus, so that the sheath can be lifted into position
around the processing component (for example the heater) as
necessary. The sheath is sized, so that when positioned in an
aperture in the cartridge, the flange (46) projects above the upper
surface of the cartridge. The sheath is made from a thermally
conducting plastics or elastomeric material, for example "CoolPoly"
materials available from Cool Polymers, Inc, thereby enabling heat
from a heater positioned within it to pass through the sheath into
the content of the chamber. The sheath preferably has a thickness
in the range of 0.25 mm to 0.75 mm its thinness assisting in heat
transfer.
[0099] A combined heater and magnet (52) used in the cartridge is
illustrated in FIGS. 11 and 12. FIG. 11 shows a side view of the
heater and magnet assembly and FIG. 12 shows the lower portion of
the assembly in cross-section. The combined heater and magnet (52)
has an elongate body, (50) with a thermally conductive (preferably
mild steel) casing (54) which houses a heating element. An assembly
body (66) is located at one end of the elongate body (50) and is
provided with a clamp (68) which clamps the elongate body (50) onto
an actuator arm (70). An insulating shim (72) separates the clamp
(68) from the elongate body (50).
[0100] A heating element is inserted into the casing (54) of the
combined heater and magnet assembly, for example as shown in FIG.
13 which is a side view of a suitable heater element (64). A
suitable heating element is one used in soldering irons, for
example a 50 Volt heating element supplied by Antex, part no.
SD50E. Such heating elements typically have a resistive wire
embedded in an insulating material, for example magnesium oxide or
a ceramic.
[0101] An integral temperature sensor (not shown) is also supplied,
enabling the temperature to be controlled. Electrical connections
(58) extend from the heating element (64) through the assembly body
(66), which provide power to the heating element (64) and
temperature sensor. The combined heater and magnet assembly may be
moved by the actuator arm (70) as illustrated by the arrow.
Alternatively, the assembly body may (66) be provided with a flange
(not shown) which is able to interact with a fork on the moveable
arm of the apparatus, to enable the combined heater and magnet to
be moved.
[0102] FIG. 12 shows a cross section of the elongate body (52) of
the combined heater and magnet. A permanent magnet (62) and heating
element (64) are located within the casing (54). The permanent
magnet (62) is located at the opposite end to the assembly body
(66). Permanent magnets are commercially available which maintain
their field strength up to over 180.degree. C. A suitable magnet,
such as an N27SH grade sintered Neodymium-Iron-Boron disc magnet is
available from MMG Mag Dev Limited. As PCR reactions are aqueous
reactions, the magnet only needs to maintain its field strength up
to 100.degree. C.
[0103] Reagents suitable for carrying out a PCR reaction may also
be prepared in a reaction chamber, for example by addition of a
suitable buffer, in particular one containing purified nucleic acid
extracted from the sample, to lyophilised beads of PCR reagents.
Again, such procedures may be effected automatically within the
apparatus by moving elements such as the cutter, pipettors etc so
as to ensure that the appropriate reagent transfers occur.
[0104] Once a PCR reaction mixture has been prepared on the
cartridge, it is suitably transferred into the reaction chamber 17,
which is thermally cyclable as a result of an ECP coating. Filling
is achieved by means of a modified pipettor and the procedure is
illustrated in FIG. 7.
[0105] As illustrated, the pipettor comprises a plastics body (25)
provided with a series of annular flanges (26) which facilitate the
collection of the pipettor by an arm of the apparatus. A cap member
(27) has a resilient upper diaphragm (28) with a projection (29)
intended to interact with an actuator provided on the apparatus, so
as to allow controlled operation of the pipettor.
[0106] The lower section (30) of the pipettor is substantially
elongate and of a sufficiently small diameter to enter a capillary
tube (31). The capillary tube (31) is sealed at the lower end (32)
and so forms a closed reaction vessel. The lower surface (32) is
suitably transparent so that the progress of any reaction carried
out in the vessel can be viewed. This means that, for example where
the PCR is carried out in the presence of a fluorescent signalling
system, it can be monitored throughout (real-time PCR). The upper
portion (33) of the reaction vessel is of a wider cross section,
but the walls in the region of the juncture of the upper portion
(33) and capillary tube (31) are tapered so as to provide a guide
for the lower section of the pipettor (30) as it enters the
capillary tube (31).
[0107] An electrically conducting polymer layer (34) surrounds the
capillary tube, and is connectable to an electrical supply by way
of upper and lower electrical contacts (35, 36).
[0108] In use, the pipettor 14 is raised out of its housing with
the cartridge by the interaction of the arm with the flanges 26,
and lowered into a reaction chamber containing the prepared PCR
reaction mixture. The pipettor actuator, driven by a stepper motor
is deployed to depress the diaphragm (28) so as to draw the
reaction mixture up into the pipettor body (25).
[0109] The pipettor is then raised out of the chamber by the
moveable arm of the apparatus, the cartridge is moved so that the
pipettor is located above the reaction vessel (17), and then
lowered, until the lower section (30) of the pipettor (14) is
substantially at the base (32) of the capillary tube.
[0110] The actuator for the diaphragm (28) is once again activated
to expel the contents into the reaction vessel (17). At the same
time, the arm is deployed to raise the pipettor (14) out of the
reaction vessel (17). The movement of the arm and the actuator are
co-ordinated so that the pipettor (14) leaves the capillary tube
(31) at a suitable rate to provide bubble free filling.
[0111] The accuracy and controllability of the actuator and the arm
as a result of the use of suitable controlling stepper motors,
means that such an operation is possible.
[0112] Once the reaction vessel (17) has been filled in this way, a
suitable cap or stopper may be applied to the upper section (33) to
close the vessel. The electrical contacts 35, 36 may be connected
so as to allow a thermal cycling process, for example a KR
reaction, to be conducted, within the reaction vessel (17) without
further movement.
[0113] Suitably, the PCR includes one of the conventional
signalling sytems such as the Taqman.TM. or ResonSense.TM.
methodologies and this is monitored through a transparent base (32)
of the tube. Once complete, the cartridge may be removed from the
apparatus and discarded.
[0114] The systems and elements described herein therefore provide
an effective and efficient way of conducting a variety of
procedures, in particular chemical, biochemical or analytical
assays, whilst minimising risks of contamination and false positive
results which this may introduce.
* * * * *