U.S. patent application number 10/496742 was filed with the patent office on 2005-04-14 for apparatus and methods for microfluidic applications.
Invention is credited to Fearnley, Joel, Ghazal, Peter, Polwart, Stuart, Roy, Douglas.
Application Number | 20050079104 10/496742 |
Document ID | / |
Family ID | 9926505 |
Filed Date | 2005-04-14 |
United States Patent
Application |
20050079104 |
Kind Code |
A1 |
Polwart, Stuart ; et
al. |
April 14, 2005 |
Apparatus and methods for microfluidic applications
Abstract
Non-rigid tape apparatus and fabrication methods for
microfluidic processing applications such as gel electrophoresis
are provided, where microfluidic processing is performed on
selected areas. Parts of the tape are formed by high pressure
plastic film forming. Membranes and other structures are self
sealing during and after penetration by pipettes and electrical
probes. Rigid exoskeleton elements protect the non-rigid parts
during processing and facilitate transport of the tape.
Inventors: |
Polwart, Stuart;
(Stirlingshire, GB) ; Fearnley, Joel;
(Peeblesshire, GB) ; Roy, Douglas; (Edinburgh,
GB) ; Ghazal, Peter; (Edinburgh, GB) |
Correspondence
Address: |
Testa Hurwitz & Thibeault
High Street Tower
125 High Street
Boston
MA
02110
US
|
Family ID: |
9926505 |
Appl. No.: |
10/496742 |
Filed: |
October 20, 2004 |
PCT Filed: |
November 27, 2002 |
PCT NO: |
PCT/GB02/05339 |
Current U.S.
Class: |
506/40 ;
422/400 |
Current CPC
Class: |
B01L 2300/021 20130101;
B01L 3/505 20130101; G01N 27/44791 20130101; B01J 2219/00317
20130101; B01L 2300/161 20130101; G01N 35/00009 20130101; Y10T
29/494 20150115; B01L 2300/042 20130101; B01J 2219/00659 20130101;
G01N 27/44782 20130101; B01J 2219/00596 20130101; B29C 51/00
20130101; B01J 2219/00495 20130101; B01L 3/502707 20130101; B01L
2300/044 20130101; B01L 2300/0812 20130101; B29C 2791/007 20130101;
B01L 2200/10 20130101; B01J 2219/00585 20130101; B01J 2219/00518
20130101; B01L 2400/0457 20130101; B01J 2219/00605 20130101; B01L
2200/12 20130101; B01L 2400/0415 20130101; B01L 2400/0406 20130101;
G01N 27/44704 20130101; C40B 60/14 20130101; B01L 2300/18 20130101;
B01J 2219/0074 20130101; B33Y 80/00 20141201; Y10T 156/1002
20150115 |
Class at
Publication: |
422/100 |
International
Class: |
G01N 001/10 |
Foreign Application Data
Date |
Code |
Application Number |
Nov 27, 2001 |
GB |
0128350.6 |
Claims
1. A microfluidic processing apparatus comprising a non-rigid
member provided with at least one microfluidic processing area and
a support member adapted to extend across the at least one
microfluidic processing area, the support member being provided
with at least one access zone for loading and/or unloading the
microfluidic processing area of the non-rigid member, wherein the
support member provides local rigidity proximate to the
microfluidic processing area of the non-rigid member.
2. The microfluidic processing apparatus of claim 1 wherein the
access zone is penetrable.
3. The microfluidic processing apparatus of claim 1 wherein the
access zone is sized to be penetrable by a sample loading
probe.
4. The microfluidic processing apparatus claim 3 wherein the sample
loading probe comprises a pipette tip.
5. The microfluidic processing apparatus of claim 1 wherein the at
least one access zone is provided with at least one access
port.
6. The microfluidic processing apparatus of claim 1 wherein the
non-rigid member is formed from a single thin polymer film and
provided with shallow channels and deep cavities on the non-rigid
member so as to provide microfluidic and macrofluidic
capabilities.
7. The microfluidic processing apparatus of claim 1 wherein
microfluidic processing can be enabled by incorporating a
conductive layers directly on to the non-rigid member.
8. The microfluidic processing apparatus of claim 7 wherein the
conductive layers are configured to act from one side of the
apparatus only, thus allowing the other side of the apparatus to be
available for other means of detecting microfluidic reaction
processes.
9. The microfluidic processing apparatus of claim 1 wherein the at
least one access zone is positioned to provide access to an end of
the microfluidic processing area.
10. The microfluidic processing apparatus of claim 1 wherein the at
least one access zone is positioned to provide access to an end
edge of the microfluidic processing area at one or more position
along its length.
11. The microfluidic processing apparatus of claim 1 wherein the
support member is affixed to the non-rigid member by means of a
snap-fit connection.
12. The microfluidic processing apparatus of claim 1 wherein the
support member is affixed to the non-rigid member by an
adhesive.
13. The microfluidic processing apparatus of claim 5 wherein the
access port is provided with a replaceable seal.
14. The microfluidic processing apparatus of claim 13 wherein the
replaceable seal is a plug.
15. The microfluidic processing apparatus of claim 1 wherein the
support member is made from a polymer film.
16. The microfluidic processing apparatus of claim 1 wherein the
support member has a thickness between 0.5 mm and 3 mm.
17. The microfluidic processing apparatus of claim 1 comprising a
plurality of support members and a plurality of microfluidic
processing areas contained on the non-rigid member, wherein one
support member extends across one microfluidic processing area.
18. The microfluidic processing apparatus of claim 1 wherein the
support member is further provided with coupling means to allow
attachment of the support member to a drive means.
19. The microfluidic processing apparatus claim 18 wherein the
coupling means comprises a plurality of apertures adapted for
engagement with the drive means.
20. The microfluidic processing apparatus of claim 1 wherein the
non-rigid member consists of a layer of deformable tape, the
microfluidic processing area thereon being formed by deformation of
the layer of deformable tape.
21. The microfluidic processing apparatus of claim 20 wherein the
deformable tape is constructed from a polymer film.
Description
[0001] This invention relates to fabrication and processing
technology for microfluidic applications in chemical and biological
processing and analysis, in particular fabrication and application
of non-rigid apparatuses optionally in the form of a tape.
[0002] In the field known as "lab-on-a-chip", electronic,
microfluidic and bio processes are combined at chip scale to bring
dramatic productivity and cost benefits to fields as diverse as
high throughput screening, bio-molecular assays and point of care
diagnostics.
[0003] Fabrication technologies are known that have been developed
in the microelectronics industry and then applied to biotechnology
and biomedical industries. However, compared to electronic based
devices, biotechnology devices are much more diverse in order to
enable the manipulation of a large variety of bio materials, fluids
and chemicals. Improvements in performance, throughput and cost
have been achieved by reducing the size and volume in miniaturised
biosystems.
[0004] These "Lab-on-a-chip" solutions have increased the amount of
functionality per apparatus by miniaturisation. The problem with
increased miniaturisation is the complexity of smaller scale
processing and the large cost of equipment for microfabrication.
Furthermore, conventional lithographic and etching processes
adopted from the microelectronics industry require rigid
apparatuses.
[0005] Glass apparatuses for microfluidic applications are known,
such as the LabCHIP from Caliper Technologies Corp (Mountain View,
Calif.), U.S. Pat. No. 6,274,089. The glass apparatus is attached
to a plastic moulded cartridge which incorporates wells for loading
test samples, reagents and-gel.
[0006] Rigid plastic apparatuses are known, such as the LabCard
from Aclara Biosciences Inc (Mountain View, Calif.), U.S. Pat. No.
6,103,199. A tooling process involving patterning and
electroplating is used to create embossed microchannels on the card
surface.
[0007] "Lab-on-a-CD" devices such as from Gamera and Gyros use
centrifugal force of a rotating disk as the micrbfluidic pumping
mechanism, e.g., Gamera Bioscience Corporation (Medford, Mass.),
U.S. Pat. No. 6,063,589.
[0008] The above are all discrete devices which require further
handling steps for continuous operation. They are also inefficient
for single test operation. Silicon apparatuses are known, such as
the Nanogen chip, which is a microfluidic microarray device, where
the microarray is selectively doped with biological or chemical
probes which can be polarised electrically to attract or repel
molecules from the sample material under test.
[0009] For example, U.S. Pat. No. 5,858,195 to Lockheed Martin
Energy Research. Corporation (Oak Ridge, Tenn.) describes a
microchip laboratory system and method to provide fluid
manipulations. The microchip is fabricated using standard
photolithographic procedures and etching, incorporating an
apparatus and rigid cover plate joined using die bonding. Capillary
electrophoresis and electrochromatography are performed in channels
formed in the apparatus. Analytes are loaded into a four-way
intersection of channels by electrokinetically pumping the analyte
through the intersection.
[0010] These approaches require time consuming additional steps of
picking and placing discrete apparatuses which increases the
overall processing cycle time in microfluidic applications.
[0011] "MicroTape.TM.--A 384 Well Ultra High Throughput Screening
System" Journal of the Association for Laboratory Automation, May
1999: Volume 4, Number 2, p. 31, Astle, T. W., teaches of a tape
device designed for storage of liquid compounds in smaller volumes
(typically 10 ul) than the industry standard 96 or 384 well
micro-titer plate (MTP). Tape storage is in a pattern identical to
a 384 well MTP In effect, MicroTape.TM. is an alternative passive
storage medium to the micro-titer plate.
[0012] The primary features of MicroTape.TM. are:
[0013] 1) bulk compounds typically stored in 96 or 384 well
micro-titer plates can be transferred into a smaller volume storage
medium, i.e. the MicroTape.TM., and then stored within the medium
for future use at low temperature. When this array of compounds is
required for test, only one section of tape (i.e. a 384 well
section) need be retrieved and defrosted, rather than the whole of
the bulk compound medium.
[0014] 2) the MicroTape.TM. incorporates a separate sealing
membrane to protect the compound during storage. This membrane is
capable of being de-sealed and re-sealed.
[0015] 3) use of MicroTape.TM. for Polymerase Chain Reaction (PCR)
processing. The concept takes a reel/roll of MicroTape.TM. and uses
alternate immersion in hot and cold water tanks to perform thermal
cycling for the PCR process.
[0016] The limitations of this approach are:
[0017] It's well capacity is 10 ul which is much larger scale than
lab-on-a-chip.
[0018] It is not patterned microfluidic channels.
[0019] It is not analytical, i.e. does not incorporate gels or
analytes through which molecular separation or purification can be
accomplished.
[0020] It is not electrically active, i.e. incorporating electrical
elements or interfacing with electrical elements i.e. it is simply
a carrier.
[0021] The PCR processing is performed on the whole reel rather
than on selectable areas or segments of the tape.
[0022] In the contemporary art of gel electrophoresis, including
the emerging field of miniaturised systems, a common means of
detection is to capture an image of these layers using
electro-optical means. A convenient method is to use a 2
dimensional CCD (Charged Coupled Device) detector array (an area
array) to capture the appearance of the permeation layer area in a
single "snapshot" image. Another convenient method is to use a 1
dimensional CCD array (a line array) and move it relative to the
permeation layer such that the full image is built up from many
adjacent line images.
[0023] It would be advantageous to provide an apparatus for
microfluidic applications that allowed an increased area for
microfluidic processing, without requiring an increase in
miniaturisation and the associated complexity of processing.
[0024] It would be further advantageous to provide an apparatus for
microfluidic applications that facilitated loading and transport of
analytes and reagents both during and after apparatus
fabrication.
[0025] It would be further advantageous to provide an apparatus
that allowed continuous processing of a moving apparatus.
[0026] It would be further advantageous to provide an apparatus
that allowed a variable area on one apparatus, while using a fixed
size of apparatus handling mechanism.
[0027] It would further be advantageous to integrate information
storage and management systems within or on the apparatus for use
with simple detection methods.
[0028] It is an object of at least one aspect of the present
invention to provide an apparatus for microfluidic
applications.
[0029] It is a further object of at least one aspect of the present
invention to allow an increased area for microfluidic processing
and novel dynamic processing steps both within and of the
apparatus, while using simple fabrication processes and apparatus
handling techniques.
[0030] In this document, a probe is defined as including mechanical
probes, electrical probes and pipettes for fluidic
manipulation.
[0031] In this document, indexing patterns are defined as including
patterns for facilitation mechanical movement, detection of
position, detection of movement, and display and recording of
information.
[0032] In this document, mass transport is defined as transport of
mass relative to the apparatus.
[0033] According to a first aspect of the present invention, there
is provided an apparatus for microfluidic processing applications,
wherein said microfluidic processing is performed on a selected
area of a plurality of areas each individually selectable on said
apparatus, characterised in that the apparatus is non-rigid.
[0034] According to a second aspect of the present invention, there
is provided an apparatus for mass transport microfluidic processing
applications, characterised in that the apparatus is non-rigid.
[0035] According to a third aspect of the present invention, there
is provided an apparatus for microfluidic processing applications,
characterised in that the apparatus comprises at least one rigid
member and at least one non-rigid member.
[0036] Preferably the apparatus comprises at least two non-rigid
members.
[0037] Preferably said non-rigid member is a tape.
[0038] Preferably there are a plurality of rigid members each
associated with one of a plurality of areas each individually
selectable on said apparatus.
[0039] Preferably said rigid member comprises access ports.
[0040] According to a fourth aspect of the present invention, there
is provided a method of fabrication of an apparatus for
microfluidic processing applications, comprising the step of
attaching at least one rigid member to at least one non-rigid
member.
[0041] Preferably said method of fabrication further comprises the
step of forming at least one non-rigid member.
[0042] Preferably said step of forming said at least one non-rigid
member comprises the step of high pressure plastic film forming
with said high pressure acting on said apparatus.
[0043] Alternatively said step of high pressure plastic film
forming is arranged with the high pressure acting on a compliant
membrane, which is part of a forming tool in contact with said
apparatus.
[0044] Preferably said rigid member has a maximum dimension
perpendicular to its plane greater than the maximum dimension
perpendicular to the plane of said at least one non-rigid
member.
[0045] According to a fifth aspect of the present invention, there
is provided a method of mounting an apparatus for microfludic
processing applications, comprising the step of attaching said
apparatus to a non-rigid carrier that is in the form of a tape.
[0046] Preferably said carrier has a maximum dimension
perpendicular to its plane greater than the maximum dimension
perpendicular to the plane of said apparatus.
[0047] Preferably said apparatus is attached to said non-rigid
carrier by snap fitting into apertures in said carrier.
[0048] Alternatively said apparatus is attached to said non-rigid
carrier by ultrasonic welding, heat sealing, adhesive, chemical or
molecular bonding.
[0049] Preferably said apparatus is a tape.
[0050] Preferably said apparatus comprises a polymer film.
[0051] Preferably said apparatus comprises processing elements for
microfluidic processing.
[0052] Typically said processing elements comprise indents of said
apparatus.
[0053] Optionally said processing elements comprise cavities
embedded within said apparatus.
[0054] Optionally said processing elements comprise processing
materials in intimate contact with the surface of said
apparatus.
[0055] Optionally said processing elements comprise processing
materials embedded within said apparatus.
[0056] Optionally said processing elements comprise opaque,
translucent or coloured materials for providing optical isolation
between elements or providing indexing marks.
[0057] Preferably an element of said apparatus is transparent.
[0058] Preferably a member of said apparatus is transparent.
[0059] Preferably said apparatus is penetrable.
[0060] Preferably said apparatus is self sealing during
penetration.
[0061] More preferably said apparatus is self sealing after
penetration.
[0062] Preferably said apparatus further comprises an impermeable
membrane.
[0063] Preferably said impermeable membrane is affixed in intimate
contact with parts of the surface of said apparatus.
[0064] Alternatively said impermeable membrane is arranged as
discrete areas of impermeable membrane in intimate contact with
parts of the surface of said apparatus.
[0065] Preferably said impermeable membrane is penetrable.
[0066] Preferably said impermeable membrane is self sealing during
penetration.
[0067] More preferably said impermeable membrane is self sealing
after penetration.
[0068] Optionally said impermeable membrane is re-sealed by a
capping element after penetration.
[0069] Preferably said impermeable membrane is supported by support
structures.
[0070] Preferably said apparatus further comprises a non-rigid
member.
[0071] Preferably said non-rigid member is affixed in intimate
contact with parts of the surface of said apparatus.
[0072] Alternatively said non-rigid member is arranged as discrete
areas of non-rigid member in intimate contact with parts of the
surface of said apparatus.
[0073] Preferably said non-rigid member is penetrable.
[0074] Preferably said non-rigid member is self sealing during
penetration.
[0075] More preferably said non-rigid member is self sealing after
penetration.
[0076] Optionally said non-rigid member is re-sealed by a capping
element after penetration.
[0077] Preferably said non-rigid member is supported by support
structures.
[0078] According to a sixth aspect of the present invention, there
is provided a method of fabrication of an apparatus for mass
transport microfluidic processing applications comprising the step
of forming an apparatus that is non-rigid.
[0079] According to a seventh aspect of the present invention,
there is provided a method of fabrication of an apparatus for mass
transport microfluidic processing applications comprising the step
of fabricating a tape.
[0080] Preferably said step of forming said apparatus comprises the
step of high pressure plastic film forming with said high pressure
acting on said apparatus.
[0081] Alternatively said step of high pressure plastic film
forming is arranged with the high pressure acting on a compliant
membrane, which is part of the forming tool in contact with said
apparatus.
[0082] Optionally said step of fabricating said apparatus further
comprises the step of preloading processing materials onto said
apparatus before fabrication.
[0083] Optionally said step of fabricating said apparatus further
comprises the step of loading processing materials onto said
apparatus during fabrication.
[0084] Typically said step of preloading or loading during
fabrication of said apparatus comprises the step of depositing
processing materials onto a carrier.
[0085] Typically said step of preloading or loading during
fabrication of said apparatus comprises the step of depositing
processing material onto a non-rigid member.
[0086] Preferably said deposited processing material comprises
permeation layers.
[0087] Alternatively said deposited processing material comprises
conductive material.
[0088] Alternatively said deposited processing material comprises
chemically or biologically active material.
[0089] Alternatively said deposited processing material comprises
marks for identity purposes.
[0090] Alternatively said deposited processing material comprises
magnetisable material.
[0091] Preferably said step of depositing comprises printing.
[0092] Alternatively said step of preloading or loading during
fabrication of said apparatus is performed by a preloading or
loading process selected from a list of processes comprising:
deposition and etching, injection into a cavity and injection into
an indentation.
[0093] Preferably said method of fabrication of said apparatus
further comprises the steps of depositing patterns on an apparatus
and forming said apparatus, wherein the localised formation of said
processing elements is responsive to the distortion by said forming
of said deposited pattern.
[0094] Preferably said method of fabrication of said apparatus
further comprises the steps of depositing patterns on an apparatus
and localised formation of said apparatus is responsive to the
topography of said deposited pattern, resulting in the formation of
said processing elements.
[0095] Preferably said step of depositing comprises
pre-printing.
[0096] According to an eighth aspect of the present invention,
there is provided a method of fabrication of an apparatus for mass
transport microfluidic processing applications, comprising the step
of including an impermeable membrane as part of said apparatus.
[0097] Preferably said step of including an impermeable membrane
comprises the step of affixing an impermeable membrane to a
substrate.
[0098] Optionally, said step of including an impermeable membrane
comprises the step of depositing, overlaying or affixing discrete
areas of impermeable membrane in intimate contact with parts of the
surface of said apparatus.
[0099] Optionally, said step of including an impermeable membrane
comprises the step of depositing, overlaying or affixing an
impermeable membrane on said apparatus and selectively removing
areas of said impermeable membrane.
[0100] Optionally, said selected removal of said impermeable
membrane is performed by the step of cropping.
[0101] According to a ninth aspect of the present invention, there
is provided a method of fabrication of an apparatus for mass
transport microfluidic processing applications, comprising the step
of including a non-rigid member as part of said apparatus.
[0102] Preferably said step of including a non-rigid member
comprises the step of affixing a non-rigid member to a
substrate.
[0103] Optionally, said step of including a non-rigid member
comprises the step of depositing, overlaying or affixing discrete
areas of non-rigid member in intimate contact with parts of the
surface of said apparatus.
[0104] Optionally, said step of including a non-rigid member
comprises the step of depositing, overlaying or affixing a
non-rigid member on said apparatus and selectively removing areas
of said non-rigid member.
[0105] Optionally, said selected removal of said non-rigid member
is performed by the-step of cropping.
[0106] According to a tenth aspect of the present invention, there
is provided a method of microfluidic processing, comprising the
steps of selecting an area of a plurality of areas of an apparatus
and performing microfluidic processing at said selected area,
characterised in that said apparatus is non-rigid.
[0107] Optionally said step of performing microfluidic processing
comprises contacting at least one conducting element that connects
the exterior of said apparatus to the interior of said
apparatus.
[0108] Preferably said method further comprises the step of
providing an electrical potential to at least one conducting
element.
[0109] Preferably said method further comprises the step of
enabling an electrical current to pass through said least one
conducting element.
[0110] Preferably said apparatus is a tape.
[0111] Preferably said microfluidic processing is mass transport
microfluidic processing.
[0112] Preferably said microfluidic processing is responsive to the
deformation of said apparatus.
[0113] Preferably said deformation comprises deformation by a step
selected from a list of steps comprising: bending, flexing,
folding, twisting, conforming to a rigid surface, mechanical
deformation, deformation by applying a sound pressure, deformation
by applying a liquid pressure, and deformation by applying a gas
pressure.
[0114] Typically said gas pressure is a negative pressure.
[0115] Optionally said deformation may further comprise the step of
bringing part of said apparatus back into contact with another part
of itself.
[0116] Alternatively, said step of deformation further comprises
the step of bringing a part of said apparatus into contact with
another apparatus.
[0117] Optionally said deformation of said apparatus comprises the
step of moving part of said apparatus into a position for
processing of said part of said apparatus.
[0118] Typically said position for processing is a position with
said apparatus in contact with a processing tool.
[0119] Preferably said microfluidic processing is responsive to
said deformation of said apparatus, said microeluidic processing
being selected from a list comprising pumping, filling, pouring,
pressurising, mixing, dispensing, aspirating, separating,
combining, heating and cooling.
[0120] According to an eleventh aspect of the present invention,
there is provided a method of processing for microfludic processing
applications, characterised in that the processing comprises the
step of piercing an impermeable membrane.
[0121] Preferably said step of piercing an impermeable membrane is
performed with at least one probe.
[0122] Optionally said at least one probe comprises at least one
pipette.
[0123] More preferably said method of processing further comprises
the step of providing an electrical potential to at least one
conducting probe that has pierced said membrane.
[0124] Alternatively said step of processing further comprises the
step of enabling an electrical current to pass through at least one
conducting probe that has pierced said membrane.
[0125] According to a twelfth aspect of the present invention,
there is provided a method of processing for microfludic processing
applications, characterised in that the processing comprises the
step of piercing an apparatus.
[0126] Preferably said apparatus is self sealing during said step
of piercing.
[0127] Preferably said apparatus is self sealing after said step of
piercing.
[0128] Optionally said apparatus is re-sealed by a capping element
after penetration.
[0129] Preferably said step of piercing the apparatus is performed
with at least one probe.
[0130] Optionally said at least one probe comprises at least one
pipette.
[0131] More preferably said method of processing further comprises
the step of providing an electrical potential to at least one
conducting probe that has pierced said apparatus.
[0132] Alternatively said step of processing further comprises the
step of enabling an electrical current to pass through a conducting
probe that has pierced said apparatus.
[0133] According to a thirteenth aspect of the present invention,
there is provided an apparatus for microfluidic processing
applications, characterised in that the apparatus is a non-rigid
tape comprising a plurality of indexing patterns.
[0134] Preferably said indexing patterns are rigid members.
[0135] Preferably said indexing patterns are repeated.
[0136] Preferably said indexing patterns are arranged to facilitate
detection of position.
[0137] Typically said indexing patterns are arranged to facilitate
detection of position using optical detection.
[0138] According to a fourteenth aspect of the present invention,
there is provided a method of transporting a tape apparatus for
microfluidic applications comprising the step of moving said
apparatus by interaction of a moving object with at least one rigid
member attached to said apparatus.
[0139] In order to provide a better understanding of the present
invention, an embodiment will now be described by way of example
only and with reference to the accompanying figures in which:
[0140] FIG. 1 illustrates in schematic form non-rigid apparatuses,
showing a section of tape and an enlargement of one area suitable
for gel electrophoresis in accordance with the present
invention;
[0141] FIG. 2 illustrates in schematic form a variety of processing
elements in accordance with the invention;
[0142] FIG. 3 illustrates processing elements incorporating
impermeable membranes comprising homogeneous apparatus
material;
[0143] FIG. 4 illustrates impermeable processing elements
incorporating discrete impermeable membranes and processing
elements on hinged tabs;
[0144] FIG. 5 illustrates the insertion and removal of a probe into
a processing element through an impermeable self-sealing
membrane;
[0145] FIG. 6 illustrates a plan view of an apparatus incorporating
an extended impermeable membrane with a variety of support
structures;
[0146] FIG. 7 illustrates a cross-section of the same structures
illustrated in FIG. 6;
[0147] FIG. 8 illustrates some of the same structures in
cross-section as FIG. 7, but where the processing elements comprise
processing materials;
[0148] FIG. 9 illustrates in schematic form a plan view of a
structure for probing through an impermeable membrane;
[0149] FIG. 10 illustrates an alternative arrangement to that of
FIG. 9 where the channel extends into the apparatus;
[0150] FIG. 11 illustrates a cross-section of the structure
illustrated in FIG. 10;
[0151] FIG. 12 illustrates a tape apparatus with indexing
patterns;
[0152] FIG. 13 illustrates in schematic form a variety of
cross-sections of indexing patterns;
[0153] FIG. 14 illustrates a flow chart describing the steps of
fabrication of an apparatus;
[0154] FIGS. 15 and 16 illustrate arrangements of scanning the
optical detectors for scanning the apparatus;
[0155] FIG. 17 illustrates plan and elevation views of a
micro-array configuration of the apparatus;
[0156] FIG. 18 illustrates in schematic form non-rigid apparatuses
in accordance with the present invention;
[0157] FIG. 19 illustrates in schematic form the components of a
planned fabrication scheme of one embodiment;
[0158] FIG. 20 illustrates in schematic form a compact fabrication
option;
[0159] FIG. 21 illustrates in schematic form an operating mode
using a vacuum suction onto a scanner or a heating/cooling
plate;
[0160] FIG. 22 illustrates in schematic form reservoir fabrication
showing the option of sample loading through penetration of a cover
seal;
[0161] FIG. 23 illustrates in schematic form reservoir fabrication
showing the option of electrical probe penetration of a cover
seal;
[0162] FIG. 24 illustrates in schematic form an alternative
electrical probe option;
[0163] FIG. 25 illustrates in schematic form a supporting layer of
one segment of a tape after preparatory printing;
[0164] FIG. 26 illustrates in schematic form a formed pattern layer
after forming;
[0165] FIG. 27 illustrates in schematic form a formed pattern layer
after a blanking operation;
[0166] FIG. 28 illustrates in schematic form a formed pattern layer
assembled to the supporting layer;
[0167] FIG. 29 illustrates in schematic form an exoskeleton;
[0168] FIG. 30 illustrates in schematic form an exoskeleton affixed
to the supporting/patterned layer;
[0169] FIG. 31 illustrates in schematic form a section (vertical
scale exaggerated for clarity) and plan view through one tape
segment and disposition of sealing plugs;
[0170] FIG. 32 illustrates in schematic form loading of electrolyte
during manufacture;
[0171] FIG. 33 illustrates in schematic form loading of analyte
during manufacture; and
[0172] FIG. 34 illustrates in schematic form loading of a test
sample at the point of use.
[0173] FIG. 35 illustrates in a flowchart of automated processing
using the fabricated tape.
[0174] The invention is a non-rigid apparatus for microfluidic
processing applications, which may be in the form of a tape. The
use of a non-rigid apparatus allows novel dynamic processing
methods. The incorporation of re-sealable impermeable layers allows
further novel dynamic processing steps.
[0175] FIG. 1a shows a typical section of tape 1 with an array of
microfluidic processing areas or processing segments 2 in
accordance with a preferred embodiment of the present invention.
Adjacent test segments are spaced to suit the sample supply vessel.
For example, where samples are delivered for test in a 384 well
microtiter plate format, the tape segments will be supplied on a
4.5 mm pitch, P. The tape is processed in a vertical plane with the
sample loading ports uppermost. The tape width, W, is typically 25
mm but is configurable in a range of 1 mm to 100 mm.
[0176] FIG. 1b shows an enlargement of a single processing segment
2, the operation of which follows well-established principles of
electrophoresis. A DNA test sample is assumed.
[0177] The apparatus includes a supporting layer 251, a formed
pattern layer 265 with a machine readable index mark 254. The
pattern layer has formed cavities 266 and a connecting channel 267
filled with gel. The exoskeleton 2915 supports plugs 3124 that are
used for re-sealable access to the cavities.
[0178] A DC voltage in the range 5 to 500 Volts (typically 100V/cm
has been found to be suitable) will be applied across negative
terminal 252 and positive terminal 253.
[0179] This will cause the negatively charged DNA sample 3430 to
migrate into the gel column 267 and its constituent molecules will
then separate into bands in accordance with their molecular weight.
An image of the band pattern will be captured by a commercial CCD
camera and the image processed and presented to the user on a
computer screen.
[0180] The electrical terminal pads 252 and 253 are conveniently
presented for perpendicular access by external contact pins whose
engagement will be controlled by the tape processing instrument.
The exoskeleton 2915 may be conveniently employed as the tape
transport means, and be driven by, for example, a toothed belt or a
drive pinion having the same tooth pitch as the test segments on
the tape.
[0181] The CCD image capture system can also conveniently capture
the test segment ID mark, thus avoiding the need for a
separate-device such as a bar code reader.
[0182] FIG. 2a illustrates a part of an apparatus 20 in
cross-section. The apparatus contains a variety of processing
elements which are an indent 21, a void or cavity in the apparatus
22 processing materials on the surface of the apparatus 23,
processing materials embedded within the apparatus 24, and
processing materials in an indent on the surface of the apparatus
25.
[0183] FIG. 2b illustrates part of an apparatus in cross-section
with processing materials partially filling the height of a cavity
in the apparatus 26 and processing material 27 embedded in a
channel 28 within the apparatus.
[0184] The processing elements may comprise geometries which have
sloping, curved or stepped surfaces. The processing materials may
be conformal layers in intimate contact with surfaces of the
apparatus. The processing elements may be opaque, translucent or
coloured in order to provide optical isolation between elements or,
alternatively, to provide indexing marks for allowing detection of
movement and position of the apparatus.
[0185] Several of the processing elements shown in FIGS. 2a and 2b
may be linked together, for example by cavities or indented
troughs, which are themselves processing elements such that the
linked elements act as a single processing group.
[0186] FIG. 2c illustrates a plan view 210 of processing element
groups 211 on part of an apparatus 212. FIG. 2d illustrates a cross
section of one of the processing element groups 211 shown in FIG.
2c. The formed plastic substrate 212 has a plastic membrane film
213 attached 214. The membrane is typically 0.1 mm thick, but could
be as thin as 0.02 mm. An indented trough 215 is provided for
processing materials such as materials based on Agarose or
polyacrylamide gel. A channel 216 is provided for a plug that can
be removed by, for example, laser ablation in order to allow
processing material transport between the indented trough 215 and
another processing element, indent 217. The substrate indents have
pips 218 that are shaped to guide a probe such as a pipette to an
area of the lower surface for penetration into the processing
elements, for example indent 217.
[0187] The substrate may be self-sealing during and after such
penetration.
[0188] The processing materials can be gases, liquids, solids or
semi-solids, e.g. biomolecular samples, fragments of DNA,
biochemical polymers, chemical polymers, biomolecular modifiers,
catalysts, antibodies, polypeptide molecules, protein molecules,
biological organisms such as cells and viruses and permeation
layers. The permeation layers may be solid, semi-solid, liquid,
viscous, gelatinous or gaseous layers. The permeation layers may be
biomolecular gates which are activated by electrical probes. The
function of the biomolecular gates is defined by their particular
depth, shape, volume and composition.
[0189] FIG. 3 shows a cross-section 30 of an apparatus for
microfluidic processing applications. The apparatus contains a
processing element 31 that is a cavity in the apparatus material.
At the top of the cavity the apparatus material is thin, such that
there is a membrane 32 that is impermeable and acts as an hermetic
seal to protect the contents of the cavity.
[0190] The apparatus contains another processing element 33, where
the membrane is configured as a flap 34, such that the cavity is
sealed when the unattached end of the membrane is in contact with
the apparatus 35.
[0191] FIG. 3 illustrates another processing element 36 with a
membrane arranged as a flap 37 and distortion of the apparatus 38
resulting in the opening of the flap at its unattached end 39.
[0192] FIG. 4a illustrates an apparatus 40 that includes the same
type of processing elements as shown in FIG. 3, but in this case
the impermeable membrane is deposited, overlaid or affixed as
discrete areas of impermeable membrane in intimate contact with
parts of the surface of the apparatus. In the first processing
element 41, the impermeable membrane 42 provides a hermetic seal to
the cavity 43.
[0193] Another processing element 44 shows the impermeable membrane
45 in intimate contact and attached to the apparatus at the left
hand side 46 and configured as a flap in a sealing contact with the
right hand side 47 of an indent in the apparatus 48. This flap may
be opened by deforming the apparatus in the same way as described
as above with reference to processing element 36.
[0194] In another processing element 49, the impermeable membrane
410 is deposited as a pluggin an indent resulting in a cavity 411,
the membrane again providing an hermetic seal.
[0195] Alternatively, the impermeable membrane is continuous with
the tape (i.e. not discrete). This continuous configuration can
also embody local flaps in the membrane and still be one continuous
membrane.
[0196] FIG. 4b illustrates a plan view and FIG. 4c illustrates
cross-section views of a strip of apparatus 413 where a section of
the apparatus had been removed 412 by punching out. The shape
punched out has left several tabs 414 each with an indent 415 for
containing processing materials. The tab 414 may be mechanically
folded along the fold line 417. The fold line may be weakened by
perforation or indenting. A second indent for processing materials
418 is positioned on the opposite side of the fold line from the
indent 415. When the tab is folded over 419, the indent 415 is
tipped over into contact with the indent 418, allowing mixing,
pouring or transfer of processing materials between the two
indents. This pouring may be assisted by the force of gravity,
capillary action or external pressure. Alternative arrangements can
be made that tilt through an angle of e.g. 30 degrees to cause
pouring.
[0197] FIG. 5 shows a cavity during a sequence of steps before
penetration 51, during penetration 52 and after penetration 53. The
probe 54, which is a pipette, is to be inserted into the cavity 55
through the membrane 56. When the probe 57 is inserted through the
membrane 58, the membrane is self-sealing, such that there is a
seal between the probe and the membrane 58. Processing materials
510 are then deposited in the cavity. After removal of the probe
511, the impermeable membrane is self-sealing and a seal 512 is
formed at the exit point of the probe. The penetration of the
impermeable membrane can allow introduction of processing materials
into cavities in the apparatus or removal of processing materials
from the apparatus, the penetration of the membrane can allow the
introduction of measurement tools into the apparatus or processing
tools into the apparatus. When penetration is by a conducting
probe, voltages can be applied that cause movement of fluids
through processing materials using an electrokinetic method.
[0198] Large areas of membrane would tend to bend on attempted
insertion of a probe. FIG. 6 shows a plan view of an apparatus 60
with an extended membrane 61 and support structures that provide
support for the membrane adjacent to the location where probes are
to penetrate the membrane. FIG. 7a shows a cross-section 70 of the
same structure that is shown in the plan view of FIG. 6. FIG. 7b
shows a cross-section 71 of the same structure that is shown in the
plan view of FIG. 6, but with a continuous membrane 72 affixed to a
substrate.
[0199] FIGS. 6 and 7 include support structures that are pillars
62, ribs 63 and an annulus 64. The centre of the annulus contains a
membrane that may be penetrated by a probe. The annulus allows a
"via" hole 65 to be created all the way through the apparatus and
through which a wire or conducting probe can be passed so that a
magnetic field can be created to interact with the adjacent
processing area of the apparatus.
[0200] Another useful structure is a circular indent but still
connected to adjacent processing elements and an externally
configured loop or coil of wire (or other conducting element)
around that circular indent. The electrical/magnetic field created
can be used to attract or trap or process the liquid in the
circular indent.
[0201] A "U" shaped pillar 66 is shown and a probe that enters in
the centre of the "U" at point 67, marked with a plus, may be
connected to a probe penetrating the impermeable membrane at the
second penetration point 68 by an electrical, liquid or permeation
path that is greater in length than the direct distance between the
two penetration points.
[0202] FIG. 8 shows a cross-section 80 of similar structures to
those in FIG. 7, except that the cavities in the apparatus are
filled with processing materials 81.
[0203] FIG. 9 shows a plan view of an apparatus 90 with a membrane
that extends from a first penetration point 91 to a second
penetration point 92 via an indented trough 93. A probe inserted
through the impermeable membrane at the first penetration point 91
may be connected to a probe penetrating the impermeable membrane at
the second penetration point 92 by an electrical, liquid or
permeation path that is greater in length than the direct distance
between the two penetration points.
[0204] FIG. 10 shows a plan view of an apparatus 100 with two
membranes, each of which are penetration points 101 and 102. The
dotted lines represent the edges of a buried channel 103 in between
the two membranes.
[0205] FIG. 11 shows a cross-section through the line connecting
the two penetration points of FIG. 10 which can be seen to be two
membranes 101 and 102. The channel 103 extends into the depth of
the apparatus 104. In this alternative arrangement the electrical,
liquid or permeation path between tips of probes that are inserted
through the penetration points are also greater than the direct
distance between the two probes.
[0206] Turning FIGS. 10 and 11 through 90 degrees, illustrates side
entry (rather than top entry) to the apparatus.
[0207] Then FIG. 10 becomes a side view of the tape and FIG. 11 is
a plan view of the plane of a strip of tape.
[0208] With reference to FIG. 12, an apparatus 120 is shown in plan
view with a plurality of indexing patterns 121. The indexing
patterns may be opaque, translucent or coloured materials. The
indexing patterns may be surface patterns, such as indents or
process materials or raised patterns of apparatus material, for
example the exoskeleton (2915 in FIGS. 1b and 29). Alternatively,
the indexing patterns may be embedded within the apparatus or
patterns of magnetism in a magnetic film or perforations through
the depth of the apparatus. Indexing patterns are arranged to
facilitate traction of the apparatus and detection of position
using optical, electromagnetic, electrochemical, electrical or
other forms of detection. The indexing patterns may also record
information-related to the apparatus processing elements or the
apparatus processing materials on the apparatus or within it
processing results, processing status, processing time, processing
location or processing identity. An indexing pattern may be a strip
of material which functions as a data recording medium, for example
magnetic or magneto-optical tape. Such tape may be written to and
read by standard methods.
[0209] With reference to FIG. 13 that shows in schematic form a
variety of cross-sections of indexing patterns, an indexing pattern
is shown as an indent 130, a raised feature 131, an embedded
feature 132 or a hole 133 punched through the apparatus.
[0210] With reference to FIG. 14a, a flow chart is shown which
describes the general process steps for the fabrication of
non-rigid apparatuses for microfluidic processing applications,
including apparatuses in the form of a tape or apparatuses of
homogeneous material which may be assembled to a tape or discrete
microfluidic devices which may be assembled to a tape.
[0211] Firstly, raw material preparation is provided, 141, the
primary material will be a flexible substrate, preferably in the
form of a continuous tape but other substrates, membranes, films,
mouldings, skeletal structures or pre-assembled microfluidic
devices may be part of the fabrication "kit".
[0212] Patterns can be pre-printed 142, preferably on a flat
plastic non-rigid substrate. These patterns may be conductive
elements, chemically or biologically active zones, magnetisable
zones, or printed marks for identity purposes.
[0213] The apparatus, 143, is formed using high pressure
thermo-forming with the high pressure acting on the apparatus or
the high pressure acting on a compliant membrane which is part of
the forming tool that is in contact with the apparatus. The high
pressure may be delivered by a gas or a fluid. During forming, the
pre-printed patterns on the tape surface may be distorted in
response to the topography of the formed processing elements. The
final position of the pre-printed pattern material may be predicted
by calibration test runs or simulation in order to design
pre-printed patterns that distort to create processing elements
that comprise the processing material that has been pre-printed.
Alternatively, the forming of an apparatus may be performed by
stereolithography or selective laser sintering. While forming the
apparatus by stereolithography or selective laser sintering,
processing elements may be included in the apparatus either by
direct patterning or in response to the topography of the
pre-printed patterns on the carrier.
[0214] The fabrication of the apparatus can further comprise the
step of preloading processing materials 144. These processing
materials may be preloaded by processes such as printing, film
deposition and etching, stereo-lithography, injecting into a cavity
and also injection into an indentation. Alternatively, the
preloading may be achieved by tilting the apparatus with respect to
gravity in order to open flaps of impermeable membrane so as to
introduce processing materials through the open flaps into
underlying structures. Alternatively these flaps may be opened by
the distortion of the apparatus, such as conforming it to a rigid
roller or corner.
[0215] A cropping operation 145 can be incorporated (optionally
before the preloading step) to insert apertures in a substrate or
finish a substrate to a defined external profile.
[0216] Apparatus assembly can continue, 146, by attachment or
assembly of other layers, for example, a sealing layer or sealing
layers, or sealing plugs, or additional supporting layers to
improve the robustness of the apparatus, or other pre-assembled
devices. The attachment methods may include a mechanical snap-fit,
a mechanical interference fit, ultrasonic welding, heat sealing,
molecular, chemical or adhesive bonding. Typically the final layer
of apparatus that is affixed results in one or more impermeable
membranes as part of the apparatus. Alternatively, the membranes
may be formed by depositing, overlaying or affixing discrete areas
of impermeable membrane in intimate contact with parts of the
surface of the apparatus. Alternatively the formation of the
impermeable membrane may be performed by depositing a film of
impermeable membrane across the apparatus and selectively removing
areas of the impermeable membrane. This selective removal may be
performed using cropping/blanking or by lithography, such as
photolithography, for patterning combined with wet or dry etching.
These membranes are optionally formed of homogeneous apparatus
material in the case of formation using stereo-lithography or
selective laser sintering.
[0217] The apparatus can incorporate a further loading sequence,
147, of chemical or biological agents such as solvents,
electrolytes, gels, stainers, dyes, affinity tags or bio-sensors.
This loading may be achieved by pipette probe through the apparatus
membrane or through an access port or access ports in the
apparatus.
[0218] These steps 141 to 147 have many possible permutations and
FIGS. 14b, 14c and 14d illustrate by way of example, the
fabrication sequence of some of the alternative embodiments
described within this document.
[0219] FIG. 14b shows the general fabrication sequence for the
three layer construction method described by FIG. 19 including the
fabrication steps 14191, 14192 and 14193 of the substrate 191
sealing layer 192 and carrier layer 193 respectively.
[0220] FIG. 14c shows the general fabrication sequence for the
three layer construction method described by FIG. 22, including the
fabrication steps 14221, 14222 and 14225 of the substrate 221
sealing layer 222 and carrier layer 225 respectively.
[0221] FIG. 14d shows the general fabrication sequence for the
construction method described by FIG. 1b including the fabrication
steps 14251, 14265, 142915 and 143124 of the substrate 251 process
layer 265, exoskeleton 2915 and sealing caps 3124 respectively.
[0222] In each of FIGS. 14a to 14d, the material preparation step
141 is a film forming step, except for the exoskeleton and sealing
cap material preparation 1411, which is a moulding step.
[0223] With reference to FIG. 15, the moving apparatus 150 with
indexing patterns that are permeation (for separation) indents 151,
can provide the scanning function of a scanning optical detector
with fixed optics 152 and a fixed line scan Charged Coupled Device
(CCD) detector 153.
[0224] Additionally, with reference to FIG. 16, when this fixed
scanning system 161 is configured to suit a chosen width of tape
apparatus 162 (e.g. 100 mm, shown in plan view, not to scale) or
multiple transverse separation layers, then it can also image
capture, without modification, any other tape apparatus which is of
lesser width 163 (e.g.50 mm or 20 mm), thus providing the advantage
of a detection system with flexibility in the handling of different
widths of substrate.
[0225] Additionally, where the substrate is configured to have more
than one discrete permeation layer in a transverse line across the
substrate, each of these more than one discrete permeation layers
can be imaged simultaneously.
[0226] In the emerging field of biological micro-arrays, the
processing substrates are typically comprised of a rigid
transparent material (e.g. a glass slide) and whereby bio-material
is deposited locally on a rectangular grid whose pitch may be in
the range of 50 um to 2 mm. The present invention provides the
advantage that it is equally suitable as a substrate for
micro-array fabrication but offers the benefit of having low
fabrication cost and a capability for continuous processing due to
the flexible nature of the apparatus in its form as a continuous
tape.
[0227] With reference to FIG. 17, the apparatus is illustrated
schematically 170 in plan and side views configured to locate each
element of a micro-array 171 in a shallow well or dimple 172, on a
tape 173, thereby allowing a reduced risk of cross contamination
between adjacent elements.
[0228] The apparatus is thus configured to provide an improved
degree of containment for any reaction process which is specified
to take place on that micro-array element and that this improved
degree of containment can allow operations of mixing, stirring or
agitation which would not be achievable with planar
micro-arrays.
[0229] The apparatus is configured such that this shallow well has
a thin wall section 174 (e.g. 0.1 mm, compared to a glass slide of
typically 1 to 3 mm) that enables the efficient coupling of a
conductive heating element 175 (for example a peltier device or
similar) to the well for the purpose of, for example, hybridisation
of a DNA sample at a temperature in the range of, for example, 60
to 80 degrees centigrade.
[0230] This thin wall section can readily be transparent and that
this enables the efficient coupling of an optical system 175 to
detect the bio-reaction state of any element on the
micro-array.
[0231] The apparatus can also have different regions functionalised
for the attachment of chemical or biological moieties such as
affinity tags or biological probes. Within a microfluidic channel,
there can be micro-zones incorporating reactive groups for highly
specific functions, e.g. an affinity tag such as a streptavidin
coated zone.
[0232] With reference to FIG. 18, an apparatus 10 according to the
present invention is shown. The apparatus 11 is non-rigid and is
shown as being bent, by the apparatus being conformed to the
surface of a roller 12.
[0233] The apparatus is non-rigid in that it is pliant, unlike
rigid apparatuses known in the prior art that are made of at least
one layer of hard plastic or glass or silicon, or where the
composite apparatus is rigid. On deformation of the apparatus
according to the present invention, the apparatus can return to its
original shape (i.e. flat) after deformation. The apparatus may
have a bend radius approaching zero.
[0234] The apparatus is a tape in that it is substantially longer
than it is wide in its larger two dimensions. Hence it is a
substantially continuous, narrow, flexible strip. The tape 13 may
be arranged in a reel-to-reel arrangement between reels or rollers
14 and 15.
[0235] With extreme deformation, the apparatus may be folded and
remain folded. This may be facilitated by using perforations or
indentations to weaken the fold line. Thus the apparatus may be
folded into a fanfold arrangement 16 for storage, dispensing and
processing.
[0236] The tape can also be separated into short discrete sections
17. The separation may be performed by guillotining or tearing
across perforations or indentations in the tape.
[0237] A continuous strip of tape 18 may be arranged around rollers
19 into a conveyor belt arrangement. A twist in the tape would
provide a Moebius strip arrangement.
[0238] The apparatus may be formed from a polymer film, that is a
thermoplastic polymer film, thermosettable polymer film,
elastomeric polymer film or hybrid compositions of each of these
films.
[0239] In another embodiment, the tape comprises three primary
construction elements as illustrated with reference to FIG. 19. The
tape incorporates a thin polymer substrate 191 that is formed to
create indented wells, channels and junctions which can be
configured to create a wide range of micro-fluidic geometries. This
substrate may optionally incorporate one or more surface coating
layers on the processing side of the substrate and these layer(s)
may fully cover the substrate surface or be confined to local areas
of the substrate. The substrate may incorporate liquid or solid
chemicals within the well or channel areas of the substrate.
[0240] The substrate and its chemical contents may be protected by
the attachment of a cover seal 192 membrane. The combined substrate
and cover seal will be attached to a carrier layer 193 whose
function is to protect the substrate from mechanical stress or
damage during handling, shipment, storage or end user processing.
The tape may be a one time use consumable item.
[0241] The tape assembly employs construction materials,
fabrication techniques and packaging methods that ensure that the
tape will function reliably at its final point of use. The tape
will therefore be unaffected by:
[0242] Automated and manual handling processes prior to shipment
packaging (factory);
[0243] Automated and manual handling processes at the point of use
(end user);
[0244] Shipment transport (protected by secondary packaging);
[0245] Transport temperatures of -40C to +70C (up to 24 hours);
[0246] Storage temperatures of 0C to +40C (up to 12 months)
[0247] Relative humidity in range 10% to 90% (transport and
storage); and
[0248] Atmospheric pressure (air cargo).
[0249] The substrate comprises a thin polymer membrane with a
thickness of 50 um preferred, but 125 um for some applications. The
thickness may be selected to match available commercial film
grades.
[0250] The substrate has:
[0251] Forming radius equal to thickness without stress
cracking;
[0252] Feature width to depth ratio, typically in range 2:1 to
1:1;
[0253] Uniform (consistent) draw during forming.
[0254] Thermal assist during (or prior to) forming is desirable.
Forming may be:
[0255] 1) high pressure in range 1 bar to 200 bar
[0256] 2) Vacuum
[0257] 3) high pressure with vacuum assistance
[0258] All of these may benefit from a pre-heating cycle.
[0259] Desirable features of the substrate include:
[0260] stable after forming (having no shape memory effects);
[0261] Flexible, non rigid, non brittle;
[0262] Abrasion Resistant;
[0263] Punchable, to create optional holes for mechanical
indexing;
[0264] Penetratable by probe (e.g. for liquid delivery or for
electrical probing);
[0265] High optical clarity;
[0266] Adaptable via suitable surface modification to minimise
static charge or to locally influence hydrophilic/hydrophobic
surface characteristics;
[0267] Chemical Resistance to Aqueous solutions
[0268] Analyte material loaded in the substrate channels typically
comprised of Agarose or Polyacrylamide,;
[0269] Provide bio-compatible surface (e.g. DNA, proteins, cells,
bacteria etc);
[0270] Avoid leeching of metals, anti-oxidants and stabilisers;
[0271] Capable of receiving a heat sealable cover layer e.g.
polyester/polyethylene cover layer; and
[0272] Printable with ink, stroke widths down to 0.1 mm.
[0273] Auxiliary coatings or deposited layers on the substrate
include:
[0274] Local conductive tracking;
[0275] Local hydrophobic coatings (e.g. PTFE);
[0276] Local hydrophilic coatings (eg titanium oxide); and
[0277] Bio-compatible coatings (e.g. parylene).
[0278] The seal 192 may be a single or composite layer but a dual
composite construction may be beneficial in that the outer layer
can be specified to resist the thermal affects of the heat sealing
tool whereas the inner layer is able to melt and create a seal
without putting the integrity of the membrane at risk. Properties
of the seal layer include:
[0279] Seal Thickness: Typically in range 10 um to 50 um;
[0280] Chemical Resistance: As per substrate above;
[0281] Optical: As per substrate above;
[0282] It is preferred that the seal be suitable for penetration by
a probe (typically 0.5-1 mm diameter) e..g. for liquid delivery or
for electrical probing. A self healing or re-sealable penetration
hole is preferred.
[0283] Pre-forming of the seal (schematically as in FIGS. 22 and
23) is optional to enhance rigidity of the sealing layer during
penetration and to provide the necessary space within the tape for
processing materials.
[0284] The carrier layer 193 can comply with EIA-481-B (Electronic
Industries Alliance), the standard for "Embossed carrier Taping"
for automated component handling in the electronic industries. A
preferred material is either black or translucent polystyrene,
preferred thickness is in the range 100 um to 300 um. This layer
will be formed prior to assembly of the substrate/cover such that
the substrate/cover will be contained within a recessed channel in
the carrier tape and thereby avoid contact with any other surfaces
during manufacture or distribution (e.g..in a reel), or at point of
use.
[0285] The primary functions of the carrier layer are a) to provide
a mechanically robust carrier for the more fragile substrate/cover
layers b) incorporate punched holes which provide a means of
transport drive for the tape c) incorporate registration features
which align the substrate/cover layer with the punched drive holes
d) incorporate apertures which allow the channels in the substrate
to be visible from underneath the tape.
[0286] With reference to FIG. 20, which is a section across the
width of the tape, not to scale, a 50 um thick microfluidic
substrate 201 formed up to 250 um deep, is contained within the 300
um thickness of the carrier 202 thus affording it protection. The
substrate has analyte 203 and is capped with the seal 204.
[0287] With reference to FIG. 21, a negative pressure (vacuum) is
applied to the two ports 210 that distorts the substrate onto a
tool 211 such as a viewing window of a scanner or a heating/cooling
plate.
[0288] With reference to FIG. 22, a sample loading probe 221 is
positioned ready to penetrate a reservoir in the pre-formed cover
seal 222 (that is dimpled for ease of insertion). The substrate
contains analyte 223 and the reservoir contains electrolyte
224.
[0289] With reference to FIG. 23, electrokinesis 231 probes are
shown penetrating the reservoirs.
[0290] With reference to FIG. 24, probes 241 external to the "wet
chemistry" zone are shown connecting to conductive layers on the
substrate that are an anode 242 and a cathode 243.
[0291] For the preferred embodiment, a single segment of tape will
be described below, comprising the means of processing one discrete
test sample of bio-material such as DNA.
[0292] FIG. 25 shows a supporting layer 251 comprises a thin flat
optically clear film of either polycarbonate, polyester,
polystyrene, poly methyl methacrylate, or other co-polymers of
these materials. This film will typically be 125 um thick but other
thicknesses in the range 25 um to 1000 um may be used. This Layer
has a pattern of conductive tracks 252 and 253 applied by screen
printing or laser printing or ink jet printing as well as a pattern
254 which can be machine read to indicate the identity of that
segment.
[0293] FIG. 26 shows a formed patterned layer 265 comprising a thin
film of either polycarbonate, polyester, polystyrene, polyethylene,
polymethyl methacrylate, polypropylene or other co-polymers of
these materials. This film will be typically 50 um thick but other
thicknesses in the range 10 um to 200 um may be used. This material
need not be optically transparent and some advantage may be gained
by having it translucent or opaque; translucency offers a means of
back-lighting scatter (opposite side from the optical supporting
layer) which may be used for illuminating and capturing an image of
the tape processes; opaqueness offers the possibility of using a
reflected front-lighting source.
[0294] High pressure thermoforming is preferably used to create
formed cavities 266, connecting channels 267, optional side
channels 268, primary access ports 269 and secondary optional
access ports 2610. Shallow channels 2611 provide entry slots for
the conductive tracks 252, 253. Typical relative depths of these
formed features is illustrated in typical section FIG. 31.
[0295] FIG. 27 shows a further preparative step in manufacturing
the formed patterned layer whereby a knifing or blanking process is
used to cut apertures or slots in the film. Apertures 2712 provide
the access entry slots for the conductive tracks 252, 253. Aperture
2713 ensures that the code mark 254 is not obscured by any
translucency or opaqueness in the film 265.
[0296] FIG. 28 shows layer 251 and layer 265 assembled together.
This will be effected by either a heat sealing or an adhesive
process or both, to ensure that the two layers achieve a tight seal
around the profile of the various patterned recesses 266, 267, 2611
etc. in Layer 265. Heat sealing can be achieved by the contact
surface material of Layer 265 comprising a thin layer of low
melting point polymer such as poly-ethylene; alternatively adhesive
bonding can comprise the use of commercial cyano-acrylate or, in
the case of sealing zones 2814, a commercial silicone rubber
compound may be used.
[0297] FIG. 29 shows an exoskeleton component 2915 whose purpose is
to protect layer 265 as well as providing rigid access ports 2916,
2917 for loading and unloading the tape. Apertures 2918 protect the
cavities 266 and an aperture 2919 protects the channel 267.
[0298] The exoskeleton material is preferably a rigid polymer such
as polycarbonate, ABS, polyester, polystyrene, polyethylene,.
polymethyl methacrylate, polypropylene or other co-polymers of
these materials. This exoskeleton will be typically 1.0 mm thick
but other thicknesses in the range 0.5 mm to 3 mm may be used.
[0299] FIG. 30 shows the rigid exoskeleton 2915 affixed to the
layer 251 plus layer 265 assembly. This may be by adhesive bonding
or by incorporating protrusions in the exoskeleton 2915 which will
snap fit into corresponding apertures in the supporting layer 251.
Where the Layer 265 adjoins an access port on the exoskeleton 2915,
for example, at cavity locations 3021, an adhesive layer,
preferably a commercial silicone rubber compound, will ensure
intimate local contact between Layer 265 and exoskeleton 2915.
[0300] FIG. 31 shows a section 3100 through the assembly 3101 along
the line "D" to "D". Depths are exaggerated in this figure for
clarity, but a typical overall height of the exoskeleton is 1 mm.
This cross section shows that cavities 266 are raised to the height
of the exoskeleton, cavities 269 are raised to a lesser extent
(typically 0.5 mm) and the channel 267 has a low profile (typically
50 to 200um deep). A conductive strip 253 (typically 20 to 50 um
thick) is shown entering a cavity 256. Sealing plugs 3124 are shown
at the access port locations. These sealing plugs will comprise
compliant polymer, preferably an elastomer such as polyurethane or
silicone rubber. These plugs will incorporate a feature allowing
removal and replacement by a simple hand tool or, for continuous
unattended operation, allow automated removal and replacement. Note
also feature 3123 which is a tapered section of cavity forming a
smooth transition between the cavity 266 and the channel 267.
[0301] FIG. 32 shows a method of loading liquid electrolyte (for
example 2 mM Tris, 2 mM Acetate, 0.5 mM EDTA) by accessing a probe
3225 into an end cavity. Locations 3226 may be vented and sealed
(plugs 3124) as part of the filling process. Note that the
micro-scale of the penetration points will allow surface tension to
prevent unwarranted leakage while the sealing caps are applied.
[0302] FIG. 33 shows a method of pre-loading a column of gel 3328
at the point of manufacture using a loading probe 3327. The gel is
loaded as a pre-determined dispensed volume from the elution cavity
end of the test segment. The gel is preloaded with a fluorescing
marker dye.
[0303] The test segment has now been pre-loaded ready for use, and
will be shipped in this condition to the point of use. The only
"wet chemistry" at the point of use is to load the test sample for
analysis.
[0304] FIG. 34 shows a loading probe 3429 penetrating through the
top loading port of the exoskeleton at the point of use. The
corresponding cap 3124 may be discarded or replaced depending on
whether the tape is required to be archived after use. The test
sample 3430 will be prepared in a solution which is denser than the
surrounding electrolyte in the tape cavity, for example, a solution
of sucrose will ensure that the test sample will flow under gravity
into the tapered channel and gather right at the top of the gel
column.
[0305] The exoskeleton incorporates access ports which can be
oriented longitudinally (e.g. port no.3431) or perpendicularly
(e.g. port no. 3432). Optionally port 3432 can be used to vent any
unwanted build up of gas in the lower cavity.
[0306] These fabrication methods can create features which provide
a wide range of processing options at the point of use.
[0307] With reference to FIG. 35, the automated processing has the
steps of transporting the tape and selecting an area for processing
3:51, piercing the apparatus with a probe 8 or probing the
apparatus 352, and performing microfluidic processing 353 at the
selected area, then repeating 354 the above steps until processing
of the reel of tape is complete.
[0308] During these steps the fabricated apparatus with its
optional preloaded processing materials may be deformed in order to
cause dynamic processing. The apparatus may be deformed by bending,
flexing, folding, twisting, conforming to a rigid surface,
mechanical deformation, deformation by applying a sound pressure,
deformation by applying a liquid-pressure, and deformation by
applying a gas pressure. Optionally the deformation can result in
the bringing of a part of the apparatus back into contact with
another part of itself or with another apparatus. The deformation
may move part of the apparatus into a position for processing,
including being in contact with a processing tool. The deformation
of the apparatus results in dynamic processing that includes
pumping, filling, pouring, pressurising, mixing, dispensing,
aspirating, separating, combining, heating and cooling.
[0309] Apparatuses that include impermeable membranes facilitate
further novel processing methods that involve the impermeable
membrane. The membrane may be pierced by one or more probes. These
probes may be pipettes.
[0310] Conducting probes that have pierced the membrane may provide
an electrical potential, and used for passing an electric current
through the conducting probe into a conducting medium.
[0311] Optionally a grid of probes are mounted on a discrete
carrier or a continuous carrier that can be indexed or replaced,
such that another set of probes can be used after the first set has
worn out.
[0312] The grid of probes may be configured such that each probe is
separately addressable and each probe may have a separate voltage
applied in order to progressively move the processing material
through processing elements, such as indented troughs and
permeation layers in the apparatus, after the grid of probes has
penetrated or contacted a corresponding grid of impermeable
membranes. This arrangement can be used to move process materials
through permeation layers for molecular separation. The controlled
and progressive switching of voltages on the grid of probes can be
used to split processing material into more than one separate
processing path through more than one separate processing elements.
These split process materials may be further combined or different
process materials may be combined at the junctions of paths through
the apparatus. In this way, the grid of electrical probes can be
configured to apply voltages that cause a multi-dimensional
separation of molecules, e.g. polypeptide or protein molecules.
[0313] If the probes are pipettes, processing materials may be
introduced into the apparatus through the impermeable membranes
that have been penetrated or processing materials removed from
within the apparatus. An array of pipettes compatible with 96, 192,
384, 1536 or 3456 well assay plates can be matched to an array of
commensurately spaced impermeable membranes for penetration by the
array of pipettes. Probes that penetrate or touch the surface of a
membrane can cause processing to be performed, such as pumping,
filling, pouring, pressurising, mixing, dispensing, aspirating,
separating, combining, heating, cooling, movement by
electrokinesis, movement by electrokinesis, movement by the
molecular entrapment method of molecular tweezers, acoustic
tweezers and bio-molecular motor principles.
[0314] An apparatus in the form of a tape may be transported
through processing equipment and handling equipment by friction of,
for example, rollers in contact with the apparatus or by pinions
inserted into indents or perforations in the apparatus in a similar
manner to the handling of photographic or cine film. Alternative
methods of moving the tape include sliding drawers and walking
beams. Moving the apparatus with electromagnetic fields and
induction within the apparatus or moving using air or fluid
pressure applied to the apparatus are also possible.
[0315] The position of the apparatus in response to movement is
detected by measurement of indexing patterns. After movement
dynamic processing can be performed and then further repeated
movement and dynamic processing steps can be performed in a
continuous fashion as the continuous tape is indexed through the
processing equipment.
[0316] In conclusion, we present the advantages of the present
invention.
[0317] A significant and long-established traditional art for some
of the kinds of bio-molecular separation described herein is
commonly referred to as "slab gel electrophoresis". The demands in
material usage, process time, operator time and workspace for this
process are recognised by those with even minor experience of this
art. The procedure commonly employs manual preparation of gels
involving mixing, heating and casting steps. Although the method
can now employ pre-cast gels to provide some degree of improvement,
the overall process remains manually intensive and inefficient.
[0318] In contrast, the present invention offers significant
advantages, by miniaturising all the elements of this traditional
process and eliminating many of the material preparation and manual
processing tasks.
[0319] While the traditional processes remain in common use, new
art is emerging which includes miniaturised bio-analysis systems
employing chip-scale technology, micro-fluidics, and semiconductor
fabrication techniques.
[0320] The present invention provides advantages over both
traditional and emerging techniques.
[0321] The present invention provides very significant savings in
materials, time and workspace over traditional gel electrophoresis
methods.
[0322] The present invention provides an adaptable platform for a
very wide range of bio-analysis processes (not just gel
electrophoresis) and employs geometric patterning, tooling methods
and fabrication methods which are much less complex than other
emerging micro-fluidic or chip scale techniques. This allows rapid
and cost effective production of multiple versions of tape to match
the range of applications anticipated.
[0323] The present invention allows bio-sample processing in a
range from one single simple test up to highly parallel and
multiple complex tests in an uninterrupted continuous serial or
parallel mode. The former is attractive to small research
laboratories, many quality control laboratories, and point of care
clinics. The latter is attractive to high throughput processing
laboratories. A combination of these processing methods is
attractive to public health hospitals and clinics whose demand can
fluctuate significantly. This range of capability is provided in
one single effective and efficient platform regardless of usage
patterns.
[0324] The present invention configures processing elements on a
highly flexible substrate and enables a versatile range of
substrate indexing patterns and transport methods to be utilised as
described.
[0325] Additionally, these transport methods provide the advantage
of allowing the use of non complex, compact, low cost optical
scanning means by the embodiment of a fixed position transverse
optical line-scanning system whose focal plane is along a line
across the width of the substrate. The scanning function is
provided by the (already provided) indexing motion of the
substrate.
[0326] This highly flexible substrate also enables the other
described features and advantages which result from bending,
folding, twisting, flexing and deforming its geometry.
[0327] The substrate flexibility also allows it to be penetrable by
probes for the purposes of processing material delivery or removal,
electrical connection and process tooling introduction.
[0328] Additionally this flexible substrate is suitable for
affixing a secondary impermeable membrane which is also readily
penetrable by suitable probes for the purposes of processing
material delivery or removal, electrical connection, process
tooling introduction.
[0329] The penetrable substrate and penetrable membrane provides a
processing system which can be fully enclosed and which can provide
some processing materials pre-loaded within the system. This
minimises preparation, avoids spillage, avoids the need for
cleaning or flushing procedures and simplifies waste disposal.
[0330] Alternatively, a stereo-lithographic method is described to
fabricate the substrate and the impermeable membrane in one
homogenous material with the advantage that this simplifies the
means of construction.
[0331] Alternatively, a selective laser sintering method is
described to fabricate the substrate and the impermeable membrane
in a single fabrication process again with the advantage that this
simplifies the means of construction.
[0332] The present invention employs one generic material type in
its construction (polymer) and avoids the significant use of glass,
silicon or metal in its fabrication. This simplifies the waste
disposal methods after bio-processing is complete.
[0333] The fabrication techniques described provide a wide range of
substrate geometries. These features can be created by rapid and
simple methods of tooling, thus avoiding the long lead times and
complexity of other miniaturised bio-processing systems.
[0334] The present invention has the advantage that these rapid and
simple fabrication techniques correspond to processing elements
whose dimensional accuracy is less critical than those of chip
scale devices. A corresponding advantage is that this is achieved
without sacrifice to the overall device size because the device
size, in the current state of the art, is determined by the
practicalities of the size of the sample loading wells and not by
the processing element sizes.
[0335] The present invention can be enhanced by pre-printing
processing materials onto a planar plastic film substrate using
commercially available printing methods and then by deforming that
substrate in a non planar fashion such that the pre-printed
material deforms into a desired shape or position and such that,
for example, a pre-printed permeation layer can subsequently (after
forming of the substrate) be hydrated into its gelatinous
phase.
[0336] Related printing and forming methods are already established
in the field of foil manufacture for "in-mould decoration" of
plastic injection moulded products (used for cosmetic effect mainly
on consumer electronic products), but the present invention
provides the scope for adapting these methods into this unconnected
field of application.
[0337] The flexible substrate is readily available in a range of
polymer materials whose optical properties can be matched to
available commercial optical systems for detection or imaging of
the bio-processing events during system operation.
[0338] Further modifications and improvements may be added without
departing from the scope of the invention herein described.
* * * * *