U.S. patent application number 10/193350 was filed with the patent office on 2003-01-02 for microfluidic systems.
This patent application is currently assigned to Caliper Technologies Corp.. Invention is credited to Bousse, Luc J., Kopf-Sill, Anne R., Parce, John Wallace.
Application Number | 20030003026 10/193350 |
Document ID | / |
Family ID | 26707203 |
Filed Date | 2003-01-02 |
United States Patent
Application |
20030003026 |
Kind Code |
A1 |
Parce, John Wallace ; et
al. |
January 2, 2003 |
Microfluidic systems
Abstract
The present invention generally provides microfluidic devices
and systems that utilize electrokinetic material transport systems
to selectively control and direct the transport of materials
through and among complex arrangements of integrated,
interconnected microscale channels disposed within integrated body
structures.
Inventors: |
Parce, John Wallace; (Palo
Alto, CA) ; Kopf-Sill, Anne R.; (Portola Valley,
CA) ; Bousse, Luc J.; (Los Altos, CA) |
Correspondence
Address: |
QUINE INTELLECTUAL PROPERTY LAW GROUP, P.C.
P O BOX 458
ALAMEDA
CA
94501
US
|
Assignee: |
Caliper Technologies Corp.
|
Family ID: |
26707203 |
Appl. No.: |
10/193350 |
Filed: |
July 10, 2002 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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10193350 |
Jul 10, 2002 |
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08971854 |
Nov 17, 1997 |
|
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6447727 |
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60031406 |
Nov 19, 1996 |
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Current U.S.
Class: |
422/400 ;
204/600; 204/601; 204/643; 422/68.1; 422/82.01; 422/82.02 |
Current CPC
Class: |
B01L 3/5025 20130101;
G01N 27/44791 20130101; B01L 3/502753 20130101; B01L 2300/0864
20130101; B01L 2400/0421 20130101; B01L 2400/0688 20130101; B01L
3/50273 20130101; B01L 2200/10 20130101; B01L 2300/0645 20130101;
B01L 2400/0418 20130101; B01L 2300/0809 20130101; Y10T 436/2575
20150115; B01L 2300/0816 20130101 |
Class at
Publication: |
422/100 ;
422/68.1; 422/82.01; 422/82.02; 422/101; 422/102; 204/600; 204/601;
204/643 |
International
Class: |
B01L 003/00; G01N
027/453 |
Claims
What is claimed is:
1. A microfluidic device, comprising: a body structure; and a
plurality of integrated microscale channels disposed in the body
structure, the plurality of integrated microscale channels
comprising: at least a first transverse channel, the first
transverse channel being in electrical communication with at least
a first electrode; at least first and second side channels disposed
on a first side of the transverse channel, each of the first and
second side channels having first and second ends, the first ends
intersecting the transverse channel, and the second ends being in
electrical communication with at least a second electrode; and
wherein an electrical current path between the at least first
electrode and the second electrode through the first side channel
provides substantially equal resistance to a resistance between the
first electrode and the second electrode through the second side
channel.
2. The microfluidic device of claim 1, further comprising: at least
third and fourth side channels disposed on a second side of the
transverse channel, each of the third and fourth side channels
having first and second ends; and wherein the first transverse
channel is in electrical communication with the first electrode via
the third and fourth side channels, the first end of the third and
fourth side channels intersecting the first transverse channel, and
the second ends of the third and fourth side channels being in
electrical communication with the at least first electrode.
3. The device of claim 1, wherein the at least first and second
side channels are in electrical communication with the first
electrode via a first channel header, the electrical resistance
between the first electrode and the transverse channel through the
first channel header and the first side channel being substantially
equal to the electrical resistance between the first electrode and
the transverse channel through the first channel header and the
second side channel.
4. The device of claim 3, wherein the first channel header has a
width that tapers as the channel header extends away from the first
electrode.
5. The device of claim 3, wherein the first channel header has a
width sufficient to provide substantially no resistance along its
length.
6. The device of claim 3, further comprising a third electrode, the
first and third electrodes being in electrical communication with
different ends of the first channel header.
7. The device of claim 6, wherein the first channel header
comprises a parabolic geometry, the first and second side channels
intersecting the first channel header at points along the first
channel header whereby a current path between the first electrode
and the transverse channel through the first channel header and the
first side channel comprises substantially equal resistance to a
current path between the third electrode and the transverse channel
through the first channel header and the second side channel.
8. The device of claim 7, wherein the current path between the
first electrode and the transverse channel through the first
channel header and the first side channel comprises substantially
the same channel length as the current path between the third
electrode and the transverse channel through the first channel
header and the second side channel.
9. The device of claim 1, wherein the at least third and fourth
side channels are in electrical communication with the second
electrode via a second channel header, the electrical resistance
between the second electrode and the transverse channel through the
second channel header and the third side channel being
substantially equal to the electrical resistance between the second
electrode and the transverse channel through the second channel
header and the fourth side channel.
10. The device of claim 9, wherein the second channel header has a
width that tapers as the channel header extends away from the
second electrode.
11. The device of claim 9, wherein the second channel header has a
width sufficient to provide substantially no resistance along its
length.
12. The device of claim 9, further comprising a fourth electrode,
the second and fourth electrodes being in electrical communication
with different ends of the second channel header.
13. The device of claim 12, wherein the second channel header
comprises a parabolic geometry, the third and fourth side channels
intersecting the second channel header at points along the second
channel header whereby a channel length between the second
electrode and the transverse channel through the second channel
header and the third side channel is substantially equal to a
channel length between the fourth electrode and the transverse
channel through the second channel header and the fourth side
channel.
14. The device of claim 1, further comprising fifth and sixth
electrodes, the fifth and sixth electrodes being in electrical
communication with different ends of the first transverse
channel.
15. The device of claim 1, wherein the first and second side
channels comprise shallow regions at their first ends, the shallow
regions having depths that are less than 50% of the depth of the
transverse channel.
16. The device of claim 1, wherein the first and second side
channels comprise shallow regions at their first ends, the shallow
regions having depths that are less than 20% of the depth of the
transverse channel.
17. The device of claim 1, wherein the first and second side
channels comprise shallow regions at their first ends, the shallow
regions having depths that are less than 10% of the depth of the
transverse channel.
18. A microfluidic device for controllably transporting material
among a plurality of intersecting microscale channels, the device
comprising: a body structure; a channel network disposed in the
body structure, the channel network comprising a plurality of
intersecting microscale channels, the plurality of intersecting
microscale channels comprising n channel intersections, and x
unintersected channel termini, wherein n is greater than or equal
to x, provided that x is at least 2 and n is at least 3; an
electrical power supply electrically coupled to each of the
unintersected channel termini, the power supply supplying a
separate electrical potential simultaneously to at least three of
the unintersected termini of the plurality of microscale channels,
the electrical potential supplied at the unintersected channel
termini controlling material transport at the n intersections.
19. The microfluidic device of claim 18, wherein n is at least
4.
20. The microfluidic device of claim 18, wherein n is at least
5.
21. The microfluidic device of claim 18, wherein n is at least
10.
22. The microfluidic device of claim 18, wherein n is at least
20.
23. The microfluidic device of claim 18, wherein x is at least
4.
24. The microfluidic device of claim 19, wherein x is at least
4.
25. The microfluidic device of claim 18, wherein x is at least
5.
26. The microfluidic device of claim 18, wherein x is at least
10.
27. The microfluidic device of claim 18, wherein x is at least
20.
28. The microfluidic device of claim 18, wherein the electrical
power supply simultaneously supplies a separate electrical
potential to at least three of the unintersected termini of the
plurality of microscale channels.
29. The microfluidic device of claim 18, wherein each of the
unintersected channel termini is in fluid communication with a port
disposed in the body structure, each port having an electrode
associated therewith, each electrode being separately and operably
coupled to the electrical power supply.
30. The microfluidic device of claim 18, wherein the plurality of
intersecting microscale channels comprises: a first transverse
channel having first and second unintersected termini; a second
transverse channel having first and second unintersected termini;
at least first and second connecting channels, each of the first
and second connecting channels having first and second ends, the
first end of the first and second connecting channels terminating
in and being in fluid communication with the first transverse
channel at first and second intersections, and the second end of
the first and second connecting channels terminating in and being
in fluid communication with the second transverse channel at third
and fourth intersections.
31. The microfluidic device of claim 18, wherein the body structure
comprises: a first substrate having a substantially planar upper
surface; at least a second substrate having a substantially planar
lower surface; and wherein the plurality of intersecting microscale
channels are fabricated as plurality of interconnected grooves into
at least one of the upper surface of the first substrate or the
lower surface of the second substrate, such that when the upper
surface of the first substrate and the lower surface of the second
substrate are mated, the plurality of interconnected grooves forms
the plurality of intersecting microscale channels.
32. A microfluidic device, comprising: a substrate having an
interconnected microscale channel network disposed therein, the
channel network comprising: a first transverse channel; at least
first and second side channels each having first and second ends,
the first and second side channels intersecting the first
transverse channel at the first ends of the first and second side
channels; at least a third side channel having a first and a second
end, the third channel intersecting the first transverse channel at
the first end of the third side channel; a first voltage source in
electrical communication with the second ends of the first and
second side channels; a second voltage source in electrical
communication with the second end of the third side channel;
wherein an electrical current path between the first voltage source
and the second voltage source via the third channel has
substantially the same electrical resistance via the first side
channel as via the second side channel.
33. A microfluidic system, comprising: a microfluidic device which
comprises: a body structure; a plurality of integrated channels
disposed in the body structure, the plurality of integrated
channels comprising: at least a first transverse channel; at least
first and second side channels disposed on a first side of the
transverse channel, each of the first and second side channels
having first and second ends, the first ends intersecting the
transverse channel, and the second ends being in fluid
communication with at least a first source of first material; at
least third and fourth side channels disposed on a second side of
the transverse channel each of the third and fourth channels having
first and second ends, the first ends being in fluid communication
with the transverse channel, and the second ends being in fluid
communication with a waste reservoir; and a material transport
system for transporting a second material into the transverse
channel, and for transporting portions of the second material into
the third and fourth channels by directing a flow of first material
from the first source, through the first and second channels into
the transverse channel.
34. A method of directing one or more materials serially introduced
into a microscale channel, into a plurality of parallel channels
fluidly connected to the microscale channel, the method comprising:
providing a microfluidic device having at least a first microscale
transverse channel, at least first and second microscale side
channels intersecting a first side of the transverse channel, at
least third and fourth microscale side channels intersecting a
second side of the transverse channel; introducing one or more
materials serially into the first transverse channel; directing at
least a portion of the one or more materials into the at least
third and fourth channels by directing material into the transverse
channel from the first and second channels.
35. A method of converting one or more materials serially
introduced into a microfluidic device into a plurality of separate
parallel channels, comprising: providing a microfluidic device that
comprises: a substrate having an interconnected microscale channel
network disposed therein, the channel network comprising: a first
transverse channel; at least first and second side channels each
having first and second ends, the first and second side channels
intersecting the first transverse channel at the first ends of the
first and second side channels; at least a third side channel
having a first and a second end, the third channel intersecting the
first transverse channel at the first end of the third side
channel; first voltage source in electrical communication with the
second ends of the first and second side channels; a second voltage
source in electrical communication with the second end of the third
side channel; wherein an electrical current path between the first
voltage source and the second voltage source via the third channel
has substantially the same electrical resistance via the first side
channel as via the second side channel; introducing the one or more
materials into the first transverse channel in a serial
orientation; and applying a current between the first voltage
source and the second voltage source to electrokinetically
transport at least a portion of the first material into each of the
first and second channels.
36. The method of claim 35, wherein in the introducing step, a
separate current is applied from each of the first and second
voltage sources to the first transverse channel to provide a
pinching flow of current from each of the first, second and third
side channels, into the first transverse channel.
37. A method of controllably transporting a material among a
plurality of interconnected microscale channels, comprising:
providing a microfluidic device having: a body structure having a
channel network disposed therein, the channel network including a
plurality of intersecting microscale channels, the plurality of
microscale channels comprising n channels and x unintersected
channel termini, wherein x is less than or equal to n, and provided
that x is at least 2 and n is at least 3; and applying a separate
selected electrical potential to at least three of the x reservoirs
simultaneously, whereby material is controllably moved at the n
intersections.
Description
CROSS-REFERENCE TO RELATED APPLICATIONS
[0001] This application claims priority to Provisional Application
No. 60/031,406, filed Nov. 19, 1996, which is hereby incorporated
herein by reference in its entirety for all purposes.
BACKGROUND OF THE INVENTION
[0002] In the electronics industry, manufacturers and developers
have sought to increase product performance, speed and capacity, as
well as the profits derived therefrom, through miniaturization.
Likewise, the pharmaceutical, biotechnology and related industries
have sought similar benefits through miniaturization and automation
of operations and processes performed in those industries.
Performance of more and more operations in less and less space has
thus become of primary interest in these industries. Space,
therefore, while perhaps not the final frontier, remains an area
that invites substantial exploitation.
[0003] To achieve this miniaturization the biotechnology and
pharmaceutical industries have recently applied some of the same
technologies which proved effective in the electronics industry,
such as photolithography, wet chemical etching, laser ablation,
etc., to the microfabrication of fluidic devices for use in
chemical and biological applications. For example, as early as
1979, researchers reported the fabrication of a miniature gas
chromatograph on a silicon wafer (discussed in Manz et al., Adv. in
Chromatog. (1993) 33:1-66, citing Terry et al., IEEE Trans.
Electron. Devices (1979) ED-26:1880). These fabrication
technologies have since been applied to the production of more
complex devices for a wider variety of applications.
[0004] There have been additional reports of microfabrication of
fluidic devices in these solid substrates for a variety of uses.
The most prominent of uses for this technology has been in the area
of microcapillary electrophoresis. Microcapillary electrophoresis
typically involves the introduction of a macromolecule containing
sample, e.g., nucleic acids or proteins, into one end of a
capillary tube that also contains a separation medium such as
agarose, polyacrylamide or the like. A potential is applied across
the capillary to draw the sample through the channel, separating
the macromolecules in the sample based upon their relative motility
in the separation medium, which can vary by the size or charge on
the macromolecules. While these methods typically employed fused
silica capillaries for the performance of electrophoretic methods,
in more recent efforts, the fused silica capillary has been
replaced by an etched channel in a solid planar substrate. A
covering substrate provides the last wall of the capillary. Early
discussions of the use of planar chip technology for fabrication of
microfluidic devices are provided in Manz et al., Trends in Anal.
Chem. (1990) 10(5):144-149 and Manz et al., Adv. in Chromatog.
(1993) 33:1-66, which describe the fabrication of fluidic devices
and particularly microcapillary devices, in silicon and glass
substrates.
[0005] The transport and direction of materials, e.g., fluids,
samples, analytes, buffers and reagents, within microfabricated
devices has generally been carried out by: (1) the incorporation of
mechanical micropumps and valves within the device (see, Published
U.K. Patent Application No. 2 248 891, Published European Patent
Application No. 568 902, U.S. Pat. Nos. 5,271,724, 5,277,556 and
5,171,132); (2) the use of electric fields to move a fluid
containing charged elements through the device (see, e.g.,
Published European Patent Application No. 376 611, Harrison et al.,
Anal. Chem. (1992) 64:1926-1932, Manz et al. J. Chromatog. (1992)
593:253-258, U.S. Pat. No. 5,126,022 to Soane); (3) the use of
acoustic energy to move fluid samples within devices by the effects
of acoustic means (see, Published PCT Application No. 94/05414 to
Northrup and White); or (4) the application of external pressure to
move fluids within the device (see, e.g., U.S. Pat. No. 5,304,487
to Wilding et al.).
[0006] As microfluidic systems become more complex, the ability to
accurately control and direct the fluid flow within these systems
becomes more and more difficult. It would therefore be desirable to
provide improved microfluidic devices or systems that take into
account the problems associated with these complex microfluidic
systems. The present invention meets these and a variety of other
needs.
SUMMARY OF THE INVENTION
[0007] The present invention is generally directed to microfluidic
systems and methods for use in performing a plurality of parallel
operations within a single microfluidic system. Such parallel
analyses may be performed on a single sample material, or upon
multiple sample materials.
[0008] In one aspect, the present invention provides a microfluidic
device, that comprises a body structure, which includes a plurality
of integrated microscale channels disposed therein. The plurality
of integrated microscale channels include at least a first
transverse channel, and at least first and second side channels
disposed on a first side of the transverse channel. Each of the
first and second side channels have first and second ends, where
the first ends intersect the transverse channel, and the second
ends are in electrical communication with at least a first
electrode. Also included are at least third and fourth side
channels disposed on a second side of the transverse channel. Each
of the third and fourth side channels similarly have first and
second ends, where the first ends intersect the transverse channel,
and the second ends are in electrical communication with at least a
second electrode. The side channels are provided whereby the
electrical current path between the first electrode and the
transverse channel through the first side channel provides
substantially equal resistance to a resistance between the first
electrode and the transverse channel through the second side
channel.
[0009] The microfluidic devices described herein are generally
useful for providing for controlled material transport within a
large number of integrated channels, with a minimum of control
nodes. For example, in a related aspect, the present invention
provides a microfluidic device for controllably transporting
material among a plurality of intersecting microscale channels. The
device comprises a body structure having a channel network disposed
therein. The channel network comprises a plurality of intersecting
microscale channels, which include n channel intersections, and x
unintersected channel termini, wherein n is greater than or equal
to x, provided that x is at least 2 and n is at least 3. An
electrical power supply is also included to supply a separate
electrical potential to each of the unintersected termini, or
electrical control nodes, of the plurality of microscale channels,
whereby the electrical potential supplied at each of the x
unintersected channel termini controls material transport at the n
intersections. In preferred aspects, the power supply utilizes a
controlled current at multiple electrodes to affect material
transport. Examples of such power supplies are described in detail
in U.S. application Ser. No. 08/678,436, and International Patent
Application No. PCT US97/12930, incorporated herein by
reference.
[0010] In an additional related aspect, the present invention
provides a microfluidic system, which includes a microfluidic
device as described above. In particular, the system includes a
microfluidic device that comprises a body structure having a
plurality of integrated channels disposed in the body structure,
the plurality of integrated channels. The integrated channels
include at least a first transverse channel, and at least first and
second side channels disposed on a first side of the transverse
channel. Each of the first and second side channels have first and
second ends, where the first ends intersect the transverse channel,
and the second ends are in fluid communication with at least a
first source of first material. Also included in the integrated
channels are at least third and fourth side channels disposed on a
second side of the transverse channel. Each of the third and fourth
channels have first and second ends, where the first ends are in
fluid communication with the transverse channel, and the second
ends are in fluid communication with a waste reservoir. The system
also includes a material transport system for transporting a second
material into the transverse channel, and for transporting portions
of the second material into the third and fourth channels. The
transport is affected by directing a flow of first material from
the first source, through the first and second channels into the
transverse channel.
[0011] The present invention also provides methods of transporting
materials in a serial to parallel material transport operation. In
particular, the present invention provides a method of directing
one or more materials serially introduced into a microscale
channel, into a plurality of parallel channels fluidly connected to
the microscale channel. The method comprises providing a
microfluidic device having at least a first microscale transverse
channel, at least first and second microscale side channels
intersecting a first side of the transverse channel, at least third
and fourth microscale side channels intersecting a second side of
the transverse channel. One or more materials are serially
introduced into the first transverse channel. At least a portion of
the one or more materials are then directed into the at least third
and fourth channels by directing material into the transverse
channel from the first and second channels.
[0012] In a further aspect, the present invention provides a method
of controllably transporting a material among a plurality of
interconnected microscale channels. The method comprises providing
a microfluidic device having a body structure that includes a
channel network disposed therein. The channel network includes a
plurality of intersecting microscale channels, which comprise n
channels and x unintersected channel termini, wherein x is less
than or equal to n, and provided that x is at least 2 and n is at
least 3. A separate selected electrical potential is applied to
each of the x reservoirs, whereupon material is controllably moved
at and through the n intersections.
BRIEF DESCRIPTION OF THE FIGURES
[0013] FIG. 1 schematically illustrates an example of multi-layer
construction of a typical microfluidic device.
[0014] FIG. 2 schematically illustrates conversion of materials in
a serial orientation in a single microscale channel into a parallel
orientation in a number of microscale channels. FIGS. 2A-2D
illustrate alternate applications of serial to parallel conversion
of sample fluids using the microfluidic devices employing the chip
layouts or designs of the present invention.
[0015] FIG. 3 illustrates an embodiment of a channel layout in a
microfluidic device for directing fluids or samples serially
introduced into the device into a number of parallel channels.
[0016] FIG. 4 illustrates a microfluidic device incorporating an
alternate channel layout for directing materials, fluids or samples
serially introduced into the microfluidic device into a plurality
of parallel channels.
[0017] FIG. 5 illustrates a microfluidic device incorporating
another alternate channel layout for directing materials, fluids or
samples serially introduced into the microfluidic device into a
large number of parallel channels.
[0018] FIG. 6 is a photograph showing the injection of separate
fluorescent material plugs (light area) into multiple parallel
channels, as schematically illustrated in FIG. 1A, in a
microfluidic device employing the geometry shown in FIG. 3.
[0019] FIG. 7 shows size based separation of nucleic acid fragments
in five parallel, interconnected channels within a single
microfluidic device.
[0020] FIGS. 8 is a schematic illustration of a device for use in
performing multiple electrophoretic separations, in parallel, of a
sample or samples introduced into the device, serially.
[0021] FIGS. 9A-C schematically illustrates a process and device
structure for converting a sample serially introduced into a
channel, into a plurality of parallel channels intersecting the
first channel.
DETAILED DESCRIPTION OF THE INVENTION
[0022] The present invention generally provides microfluidic
devices and systems that utilize electrokinetic material transport
systems to selectively control and direct the transport of
materials through and among complex arrangements of integrated,
interconnected microscale channels disposed within integrated body
structures.
[0023] As used herein, the term "microscale" or "microfabricated"
generally refers to structural elements or features of a device
which have at least one fabricated dimension in the range of from
about 0.1 .mu.m to about 500 .mu.m. Thus, a device referred to as
being microfabricated or microscale will include at least one
structural element or feature having such a dimension. When used to
describe a fluidic element, such as a passage, chamber or conduit,
the terms "microscale," "microfabricated" or "microfluidic"
generally refer to one or more fluid passages, chambers or conduits
which have at least one internal cross-sectional dimension, e.g.,
depth, width, length, diameter, etc., that is less than 500 .mu.m,
and typically between about 0.1 .mu.m and about 500 .mu.m. In the
devices of the present invention, the microscale channels or
chambers preferably have at least one cross-sectional dimension
between about 0.1 .mu.m and 200 .mu.m, more preferably between
about 0.1 .mu.m and 100 .mu.m, and often between about 0.1 .mu.m
and 20 .mu.m. Accordingly, the microfluidic devices or systems
prepared in accordance with the present invention typically include
at least one microscale channel, usually at least two intersecting
microscale channels, and often, three or more intersecting channels
disposed within a single body structure. Channel intersections may
exist in a number of formats, including cross intersections, "T"
intersections, or any number of other structures whereby two
channels are in fluid communication.
[0024] The body structure of the microfluidic devices described
herein typically comprises an aggregation of two or more separate
layers which when appropriately mated or joined together, form the
microfluidic device of the invention, e.g., containing the channels
and/or chambers described herein. Typically, the microfluidic
devices described herein will comprise a top portion, a bottom
portion, and an interior portion, wherein the interior portion
substantially defines the channels and chambers of the device.
[0025] In preferred aspects, the microfluidic devices include
multi-layer body structures in which the microscale channels are
disposed, one example of which is illustrated in FIG. 1. The bottom
portion of the device 12 comprises a solid substrate that is
substantially planar in structure, and which has at least one
substantially flat upper surface 14. A variety of substrate
materials may be employed as the bottom portion. Typically, because
the devices are microfabricated, substrate materials will be
selected based upon their compatibility with known microfabrication
techniques, e.g., photolithography, wet chemical etching, laser
ablation, air abrasion techniques, LIGA methods, injection molding,
embossing, and other techniques. The substrate materials are also
generally selected for their compatibility with the full range of
conditions to which the microfluidic devices may be exposed,
including extremes of pH, temperature, salt concentration, and
application of electric fields. Accordingly, in some preferred
aspects, the substrate material may include materials normally
associated with the semiconductor industry in which such
microfabrication techniques are regularly employed, including,
e.g., silica based substrates, such as glass, quartz, silicon or
polysilicon, as well as other substrate materials, such as gallium
arsenide and the like. In the case of semiconductive materials, it
will often be desirable to provide an insulating coating or layer,
e.g., silicon oxide, over the substrate material, and particularly
in those applications where electric fields are to be applied to
the device or its contents. The production of microfluidic devices
according to the present invention, uses for such devices, methods
of operating such devices, and peripheral devices for use with such
microfluidic devices, are generally described in U.S. patent
application Ser. Nos. 08/691,632, 08/683,080, 08/671,987,
08/678,436 and provisional U.S. Patent Application Serial No.
60/015,498, each of which is hereby incorporated herein by
reference in its entirety for all purposes.
[0026] In additional preferred aspects, the substrate materials
comprise polymeric materials, e.g., plastics, such as
polymethylmethacrylate (PMMA), polycarbonate,
polytetrafluoroethylene (TEFLON.TM.), polyvinylchloride (PVC),
polydimethylsiloxane (PDMS), polysulfone, and the like. Such
polymeric substrates are readily manufactured using available
microfabrication techniques, as described above, or from
microfabricated masters, using well known molding techniques, such
as injection molding, embossing or stamping, or by polymerizing the
polymeric precursor material within the mold (See U.S. Pat. No.
5,512,131). Such polymeric substrate materials are preferred for
their ease of manufacture, low cost and disposability, as well as
their general inertness to most extreme reaction conditions. Again,
these polymeric materials often include treated surfaces, e.g.,
derivatized or coated surfaces, to enhance their utility in the
microfluidic system, or may be selected so as to provide an
appropriate surface charge, e.g., provide enhanced fluid direction,
e.g., as described in U.S. patent application Ser. No. 08/843,212,
filed Apr. 14, 1997 (Attorney Docket No. 17646-002610), and which
is incorporated herein by reference in its entirety for all
purposes.
[0027] The channels and/or chambers of the microfluidic devices are
typically fabricated into the upper surface of the bottom substrate
or portion 12, as microscale grooves or indentations 16, using the
above described microfabrication techniques. The top portion or
substrate 18 also comprises a first planar surface 20, and a second
surface 22 opposite the first planar surface 20. In the
microfluidic devices prepared in accordance with the methods
described herein, the top portion also includes a plurality of
apertures, holes or ports 24 disposed therethrough, e.g., from the
first planar surface 20 to the second surface 22 opposite the first
planar surface.
[0028] The first planar surface 20 of the top substrate 18 is then
mated, e.g., placed into contact with, and bonded to the planar
surface 14 of the bottom substrate 12, covering and sealing the
grooves and/or indentations 16 in the surface of the bottom
substrate, to form the channels and/or chambers (i.e., the interior
portion) of the device at the interface of these two components.
The holes 24 in the top portion of the device are oriented such
that they are in communication with at least one of the channels
and/or chambers formed in the interior portion of the device from
the grooves or indentations in the bottom substrate. In the
completed device, these holes function as reservoirs for
facilitating fluid or material introduction into the channels or
chambers of the interior portion of the device, as well as
providing ports at which electrodes may be placed into contact with
fluids within the device, allowing application of electric fields
along the channels of the device to control and direct fluid
transport within the device.
[0029] In many embodiments, the microfluidic devices will include
an optical detection window disposed across one or more channels
and/or chambers of the device. Optical detection windows are
typically transparent such that they are capable of transmitting an
optical signal from the channel/chamber over which they are
disposed. Optical detection windows may merely be a region of a
transparent cover layer, e.g., where the cover layer is glass or
quartz, or a transparent polymer material, e.g., PMMA,
polycarbonate, etc. Alternatively, where opaque substrates are used
in manufacturing the devices, transparent detection windows
fabricated from the above materials may be separately manufactured
into the device.
[0030] These devices may be used in a variety of applications,
including, e.g., the performance of high throughput screening
assays in drug discovery, immunoassays, diagnostics, genetic
analysis, and the like. As such, the devices described herein, will
often include multiple sample introduction ports or reservoirs, for
the parallel or serial introduction and analysis of multiple
samples. Alternatively, these devices may be coupled to a sample
introduction port, e.g., a pipettor, which serially introduces
multiple samples into the device for analysis. Examples of such
sample introduction systems are described in e.g., U.S. patent
application Ser. Nos. 08/761,575 and 08/760,446 (Attorney Docket
Nos. 17646-000410 and 17646-000510, respectively) each of which was
filed on Dec. 6, 1996, and is hereby incorporated by reference in
its entirety for all purposes.
[0031] As noted previously, the devices, methods and systems
described herein, employ electrokinetic material transport systems,
and preferably, controlled electrokinetic material transport
systems to controllably direct materials among the various channels
contained within the device. As used herein, "electrokinetic
material transport systems" include systems which transport and
direct materials within an interconnected channel and/or chamber
containing structure, through the application of electrical fields
to the materials, thereby causing material movement through and
among the channel and/or chambers.
[0032] Such electrokinetic material transport and direction systems
include those systems that rely upon the electrophoretic mobility
of charged species within the electric field applied to the
structure. Such systems are more particularly referred to as
electrophoretic material transport systems. Other electrokinetic
material direction and transport systems rely upon the
electroosmotic flow of fluid and material within a channel or
chamber structure, which results from the application of an
electric field across such structures. In brief, when a fluid is
placed into a channel which has a surface bearing charged
functional groups, e.g., hydroxyl groups in etched glass channels
or glass microcapillaries, those groups can ionize. In the case of
hydroxyl functional groups, this ionization, e.g., at neutral pH,
results in the release of protons from the surface and into the
fluid, creating a concentration of protons at near the
fluid/surface interface, or a positively charged sheath surrounding
the bulk fluid in the channel. Application of a voltage gradient
across the length of the channel, will cause the proton sheath to
move in the direction of the voltage drop, i.e., toward the
negative electrode.
[0033] "Controlled electrokinetic material transport and
direction," as used herein, refers to electrokinetic systems as
described above, which employ active control of the voltages
applied at multiple, i.e., more than two, electrodes. Rephrased,
such controlled electrokinetic systems concomitantly regulate
voltage gradients applied across at least two intersecting
channels. Controlled electrokinetic material transport is described
in Published PCT Application No. WO 96/04547, to Ramsey, which is
incorporated herein by reference in its entirety for all purposes.
In particular, the preferred microfluidic devices and systems
described herein, include a body structure which includes at least
two intersecting channels or fluid conduits, e.g., interconnected,
enclosed chambers, which channels include at least three
unintersected termini. The intersection of two channels refers to a
point at which two or more channels are in fluid communication with
each other, and encompasses "T" intersections, cross intersections,
"wagon wheel" intersections of multiple channels, or any other
channel geometry where two or more channels are in such fluid
communication. An unintersected terminus of a channel is a point at
which a channel terminates not as a result of that channel's
intersection with another channel, e.g., a "T" intersection. In
preferred aspects, the devices will include at least three
intersecting channels having at least four unintersected termini.
In a basic cross channel structure, where a single horizontal
channel is intersected and crossed by a single vertical channel,
controlled electrokinetic material transport operates to
controllably direct material flow through the intersection, by
providing constraining flows from the other channels at the
intersection. For example, assuming one was desirous of
transporting a first material through the horizontal channel, e.g.,
from left to right, across the intersection with the vertical
channel. Simple electrokinetic material flow of this material
across the intersection could be accomplished by applying a voltage
gradient across the length of the horizontal channel, i.e.,
applying a first voltage to the left terminus of this channel, and
a second, lower voltage to the right terminus of this channel, or
by grounding the right terminus. However, this type of material
flow through the intersection would result in a substantial amount
of diffusion at the intersection, resulting from both the natural
diffusive properties of the material being transported in the
medium used, as well as convective effects at the intersection.
[0034] In controlled electrokinetic material transport, the
material being transported across the intersection is constrained
by low level flow from the side channels, e.g., the top and bottom
channels. This controlled material transport at the intersections
is accomplished by applying a slight voltage gradient along the
path of material flow, e.g., from the top and bottom termini of the
vertical channel, toward the right terminus. The result is a
"pinching" of the material flow at the intersection, which prevents
the diffusion of the material into the vertical channel. The
pinched volume of material at the intersection may then be injected
into the vertical channel by applying a voltage gradient across the
length of the vertical channel, i.e., from the top terminus to the
bottom terminus. In order to avoid any bleeding over of material
from the horizontal channel during this injection, a low level of
flow is directed back into the side channels, resulting in a "pull
back" of the material from the intersection.
[0035] In addition to pinched injection schemes, controlled
electrokinetic material transport is readily utilized to create
virtual valves, which include no mechanical or moving parts.
Specifically, with reference to the cross intersection described
above, flow of material from one channel segment to another, e.g.,
the left arm to the right arm of the horizontal channel, can be
efficiently regulated, stopped and reinitiated, by a controlled
flow from the vertical channel, e.g., from the bottom arm to the
top arm of the vertical channel. Specifically, in the `off` mode,
the material is transported from the left arm, through the
intersection and into the top arm by applying a voltage gradient
across the left and top termini. A constraining flow is directed
from the bottom arm to the top arm by applying a similar voltage
gradient along this path (from the bottom terminus to the top
terminus). Metered amounts of material are then dispensed from the
left arm into the right arm of the horizontal channel by switching
the applied voltage gradient from left to top, to left to right.
The amount of time and the voltage gradient applied dictates the
amount of material that will be dispensed in this manner. Each of
the above material flow profiles, e.g., pinched, gated or
pull-back, are examples of controlled material transport at
intersections.
[0036] Controlled electrokinetic material transport at
intersections also permits/encompasses relatively precise mixing of
materials from two channels that meet at a common intersection
using electrokinetic material transport systems. Specifically, by
applying appropriate electrokinetic forces to each channel, e.g.,
at a desired ratio, one can dictate relatively precisely, the ratio
of materials being mixed from each of the channels.
[0037] Although described for the purposes of illustration with
respect to a single four-way, cross intersection, in accordance
with the present invention, such systems are readily adapted for
more complex channel geometries. For example, as set forth in U.S.
patent application Ser. Nos. 08/671,987 and 08/671,986, both filed
Jun. 28, 1996 and incorporated herein by reference, microfluidic
devices are often utilized to perform a large number of parallel
operations on a sample or a number of samples, i.e., to screen
biological samples, to screen test compounds for drug discovery,
and the like. To carry out these operations, a substrate will
typically employ an array of parallel channels interconnected by
one or more common channels. Fluids required for the subject
reaction, e.g., samples or reagents, are directed along one or more
of the common channels, and are delivered to each of the parallel
channels. The materials must then be converted from the serial
orientation in the common channel, into a parallel orientation in
the various parallel channels. The present invention provides
devices, systems and methods for accomplishing this conversion.
[0038] In one aspect, the present invention generally provides
novel substrate channel layouts that ensure controlled material
transport in interconnected parallel channels, i.e., connected to a
common transverse channel, and thereby facilitates the direction of
fluids or samples serially introduced into a microfluidic device,
into a number of separate parallel channels. The direction of
materials from a serial orientation in one channel, to a parallel
orientation in a plurality of channels within these microfluidic
devices is generally referred to herein as "serial to parallel
conversion."
[0039] Serial to parallel conversion of materials within a
microfluidic device is important for a number of reasons. For
example, where one is performing a number of separate analyses on a
single sample, serial to parallel conversion can be used to aliquot
the single sample among a number of separate channels in a
microfluidic device, wherein a different analysis or assay is
performed in each different channel. Alternatively, a number of
physically discrete and different samples, e.g., drug candidates,
diagnostic samples, or the like, may be serially introduced into a
single device and allocated among a number of different parallel
channels subjecting the samples to the same basic analysis.
[0040] Schematic illustrations of serial to parallel conversions
are shown in FIGS. 2A-2D. For example, in FIG. 2A, a single plug of
sample material (1) is shown being converted to a plurality of
separate aliquots of the sample material, in a series of parallel
channels. Alternatively, as shown in FIG. 2B, separate aliquots of
the same sample material, provided in a serial orientation in a
single channel are allocated to each of several separate parallel
channels. In a particularly useful aspect, as shown in FIG. 2C, a
plurality of different compounds (1, 2, 3 and 4) are serially
introduced into a microscale channel (top) and then are each
redirected to a separate parallel channel for separate analysis.
FIG. 2D also illustrates a particularly useful application of
serial to parallel conversion where a plurality of different
samples (1, 2, 3 and 4) are serially introduced into a microfluidic
channel, and are allocated and redirected among a number of
parallel channels, wherein each parallel channel receives a portion
of each of the samples and reflects the serial orientation
originally presented (bottom). This is a particularly useful
application in the ultra high throughput analysis of large numbers
of sample materials, e.g., where a plurality of different samples
(e.g., 1, 2, 3 and 4) may be subjected to a plurality of different
analyses (e.g., in each separate parallel channel).
[0041] While serial to parallel conversion is an important aspect
of fluid control in microfluidic systems, it is not without its
problems. For example, as noted above, material transport in
electrokinetic systems is driven by current flow between electrodes
disposed at different points in the system. Furthermore, resistance
in the fluid channels, which is inversely related to current flow,
changes as a function of path length and width, and thus, different
length channels will have different resistances. In systems having
multiple parallel current paths, e.g., channels, interconnected to
a common channel, material transport along each of the parallel
channels is most easily controlled by providing electrode pairs at
the termini of each of the parallel channels. These pairs of
electrodes are then used to provide matching currents in each of
the parallel channels. Specifically, use of common electrodes for
all of the parallel channels, without more, can result in the
formation of transverse electric fields among the various parallel
channels. These transverse electrical fields can destroy the
ability of the devices to direct fluid flow within these devices.
Specifically, the current, and thus the fluid flow, will follow the
path of least resistance, e.g., the shortest path, between
electrodes. By presenting matching currents in these parallel
channels, one avoids the formation of these transverse electrical
fields from one parallel channel to the next. While the use of
separate pairs of electrodes for each channel will obviate the
problem of transverse electrical fields, production of devices
incorporating this many electrodes, and control systems for
controlling the electrical potential applied at each of these
electrodes would prove prohibitively complex. This is particularly
true where one is dealing with tens, hundreds or even thousands of
parallel channels in a single small-scale device, e.g., 1-10
cm.sup.2. Accordingly, the present invention provides microfluidic
devices for affecting serial to parallel conversion, by ensuring
that current flow through each of a plurality of parallel channels
is either equal, or is at some preselected level, without requiring
separate electrodes for each separate parallel channel.
[0042] Generally, the devices and systems of the present invention
accomplish this controlled electrokinetic transport in
interconnected parallel channel structures by ensuring that the
current flow through each of the parallel channels is substantially
equal. Maintaining substantially equal current flow is generally
accomplished by controlling the amount of resistance along any
given current path. This, in turn, may be accomplished by
controlling the path length or width, or a combination of the two,
at least over a portion of the current path.
[0043] In addition to permitting control of materials in
interconnected parallel channel networks without the formation of
transverse electric fields, as noted above, the present invention
also provides the advantage of permitting control of material
transport in a large number of interconnected channels, and at a
plurality of channel intersections, with a minimum number of
electrical control nodes in the fluidic system (e.g.,
electrodes).
[0044] By "control of material transport" at an intersection is
meant the precise control of material direction and flow rates from
the channel segments into the intersection with which these
segments are in fluid communication. Typically, such controlled
material transport utilizes the controlled input of current into
the intersection from the various channel segments that communicate
with that intersection. Such precise control is used to precisely
control the flow of material into the intersection from each of the
channel segments, or alternatively, to prevent material from
flowing into the intersection from one or more channel segmennts,
through the control of current at the unintersected channel termini
in the overall channel network. In the first illustrated example,
controlled material transport at an intersection involves the
simultaneous application of current flow from at least two channel
segments into a particular intersection. Thus, in this respect,
controlled material transport at an intersection involves more than
unidirectional flow of material through an intersection, e.g., from
one channel segment into the intersection and out through another
channel segment. Instead, material is transported into the
intersection through at least two channel segments, and out through
a third. Examples of this type of controlled transport at an
intersection include, e.g., mixing flow, pinched flow, gated flow
and/or `pull-back` flow of material.
[0045] Control of material transport at an intersection also
utilizes precise current control to prevent materials from being
flowed into a given intersection. This type of control is
schematically illustrated in a parallel channel structure, e.g., as
shown in FIGS. 9A-9C. As shown in FIG. 9A, the device is
schematically illustrated having a central sample loading channel
(connecting reservoirs 3 and 4), and two linking channels
(connecting reservoirs 1 and 2, and 5 and 6, respectively) also
termed transverse channels. Connecting each of the linking channels
and intersecting the sample loading channel in the process are a
number of separation channels. As described in substantially
greater detail, herein, the separation channels, typically in
combination with the linking channels are designed to provide
equivalent electrical resistance between electrodes, regardless of
the parallel channel used.
[0046] In operation, sample(s) are loaded through the sample
loading channel by applying a current through the sample loading
channel. In order to prevent current looping through the separation
channels and linking channels, equivalent currents are applied
through the linking channels, such that the net current through the
separation channels is zero (FIG. 9B). Relative applied voltages
are indicated for each of the reservoir/electrodes, e.g., V1, V2,
etc. Pinched flow in the sample loading channel can also be
provided by applying a slightly smaller voltage gradient across the
linking channels, causing a very slight level of current flow
through the separation channels from the linking channels into the
sample loading channel. For example, a 10 mA current can be applied
to reservoir 3. A pinching flow is applied by applying 10 mA at
reservoirs 1 and 5 and only taking out 9.9 mA at reservoirs 2 and
6. The remaining 0.1 mA forces flow toward the sample loading
channels by taking off 10.2 mA at reservoir 4.
[0047] Sample(s) disposed across the intersection of the sample
loading channel and the separation channels is/are then injected
into each of the separation channels and separated by applying the
current from reservoirs 1 and 2 to reservoirs 5 and 6 (FIG.
9C).
[0048] This same operation is illustrated with respect to the
device shown in FIG. 3, as follows. In brief, a material is
transported through channel 202 across that channel's intersections
with channels 208-244, by applying a current through that channel,
e.g., a voltage gradient between reservoirs 204 and 206. Material
transport at these intersections is controlled by applying a
matching current through channels 224 and 246, thereby preventing
current flow through any of the side channels 208-244. This is
described in greater detail, below.
[0049] Thus, control of material transport at an intersection is
characterized by the ability to control the level of current flow
into an intersection from each of the channel segments that
communicate with that intersection, including preventing current
flow from or into some of those channel segments.
[0050] As used herein, the term "material" generally refers to
molecular species that are typically fluid borne. For example, in
systems utilizing electroosmotic material transport, the material
of interest typically includes the bulk fluid and all of its
constituent elements that are being transported. In electrophoretic
material transport systems, however, the material that is being
transported includes charged molecular species, including the
material of interest, e.g., sample components, as well as ionic
species that are moving under such electrophoretic control, e.g.,
buffer salts, ions and the like.
[0051] Typically, control of material transport at a particular
intersection by electrokinetic means, requires a separate electrode
at the unintersected terminus of each channel that communicates at
the intersection. Thus, where a microfluidic system has a single
intersection made up of four intersecting channels (a typical
four-way or crossing intersection), four electrodes, typically
disposed at the unintersected termini of the four channels, would
be required to control material movement from each channel into the
intersection. Similarly, in a system having a simple "T"
intersection where three channel segments communicate at the
intersection, control of material transport at the intersection
would require at least three separate electrical control nodes,
typically disposed at the unintersected termini of the channel
segments. Furthermore additional added intersections typically
require the addition of at least one new electrode to control
material transport at the new intersection. Specifically, a channel
network made up of a main channel intersected at two different
points by two separate channels, e.g., two "T" intersections,
requires four separate electrodes disposed at the unintersected
termini of the channel segments.
[0052] Based upon the foregoing, it can be seen that microfluidic
systems having multiple intersections typically required a large
number of electrodes to control material transport at those
intersections. Specifically, a system having n intersections would
typically require at least (n+2) electrodes, to control material
transport at those intersections (assuming the simplest geometry of
a main channel intersected by multiple other channels at multiple
"T" intersections). For more complex systems, e.g., parallel
channels disposed between two common, transverse channels, even
more electrodes are required, e.g., n+4 electrodes. Rephrased, in
typically described microfluidic systems, the number of
intersections is always less than, and often, far less than the
number of electrodes used to control material transport at those
intersections.
[0053] The microfluidic devices and systems of the present
invention, on the other hand, include a plurality of intersecting
microscale channels that include at least n channel intersections,
and x electrical control nodes at the unintersected channel
termini, as described above. In these devices, however, the number
of channel intersections (n) is always greater than or equal to the
number of electrical control nodes disposed at unintersected
channel termini (x), provided that there are at least 2 channel
intersections, preferably, at least 3 channel intersections, and at
least 2 electrical control nodes. The devices of the present
invention optionally include at least 4, 5, 10 or even 20 or more
channel intersections. Accordingly, the devices of the present
invention optionally include at least 3, 4, 5, 10 or even 20
different electrical control nodes.
[0054] By way of example, a microfluidic device that incorporates
10 parallel channels connecting two transverse channels includes 20
intersections, 1 intersection where each parallel channel (10)
intersects each transverse channel (2). Material transport at these
20 intersections can be controlled according to the present
invention, by simply controlling the potentials applied at the
termini of the transverse channels, of which there are 4. This
compares favorably to the use of electrodes at the termini of each
parallel channel to control material transport at the intersection
of these channels with the transverse channels. In particular, such
a system in the example provided, would require the four electrodes
described above, as well as an additional 20 electrodes at each
parallel channel terminus, for a total of 24 or (n+4)
electrodes.
[0055] For purposes of clarification, as used herein, the term
"intersection" refers to a point in a microfluidic channel system
or network at which three or more channels or channel segments are
in fluid communication. Thus, as alluded to above, an intersection
includes a simple "T" intersection, at which three channel segments
communicate, as well as a simple cross or four-way intersection.
Other types of intersections are also included within this
definition, including, e.g., radial intersections, also termed
"wagon wheel" intersections, at which larger numbers of channel
segments, e.g., five or more, are in fluid communication.
[0056] A number of different channel geometries and layouts can be
utilized to provide substantially equal currents in interconnected
parallel channels, while requiring a minimum number of electrical
control nodes. Several of these geometries are illustrated in FIGS.
3-5 and FIG. 8 For example, in one embodiment, FIG. 3 illustrates a
microfluidic device fabricated from a planar substrate 200. The
device employs a channel orientation that may be used to accomplish
serial to parallel conversion or equal fluid flow in parallel
channels. The substrate 200 includes main channel 202, which
includes electrodes disposed in each of ports 204 and 206, at the
termini of channel 202. A series of parallel channels 208-222 and
230-244 terminate in main channel 202. The opposite termini of
these parallel channels are connected to parabolic channels 224 and
246, respectively. Electrodes are disposed in ports 226, 228, 248
and 250, which are included at the termini of these parabolic
channels, respectively. Thus, in the device shown, material
transport at the intersections of channels 208-222 and 230-244 with
parabolic channels 246 and 224, respectively, as well as transverse
channel 202, is controlled by application of appropriate voltages
at reservoirs 204, 206, 226, 228, 248 and 250. Thus, application of
voltages at 6 reservoirs controls material transport and direction
at 32 different channel intersections. The overall device shown
includes 33 channel intersections and only 8 reservoirs at which
voltages are applied, also termed electrical control nodes.
[0057] In operation, a fluid or sample plug is pumped along main
channel 202 by applying a voltage gradient between electrodes 204
and 206. An equal voltage gradient is applied between electrodes
226 and 228, and 248 and 250, resulting in a net zero flow through
the parallel channels. Specifically, no voltage gradient exists
along the length of these parallel channels.
[0058] The sample may be present within main channel 202 as a long
slug of a single sample, or multiple slugs of a single or multiple
samples. Once the sample material or materials reach the
intersection of the main channel with the parallel channels, e.g.,
230-244, it is then directed into and through the parallel channels
by applying a potential gradient between electrodes 226:246, and
228:248, which results in a material transport from parallel
channels 208-222, to force the samples into parallel channels
230-244. The resistance, and thus the current flow, in each of the
parallel channels 208-222 and 230-244 is maintained constant by
adjusting the length of the parallel channels, resulting in a
parabolic channel structure connecting each of the parallel
channels to its respective electrodes. The resistance within the
parabolic channel between parallel channels is maintained constant
by adjusting the channel width to accommodate variations in channel
length resulting from the parabolic shape of the overall channel.
For example, channel segment 224a, while longer than channel
segment 224b, will have the same resistance, because segment 224a
is appropriately wider. Thus, the parabolic design of channels 224
and 246, in combination with their tapering structures, results in
the resistance along all of the parallel channels being equal,
resulting in an equal fluid flow, regardless of the path chosen.
Generally, determining the dimensions of channels to ensure that
the resistances are equal among the channels, may be carried out by
well known methods, and generally depends upon factors such as the
make-up of the fluids being moved through the substrates.
[0059] In another example, FIG. 4 illustrates how the principles of
the present invention can be used in a substrate design that
employs fewer electrodes to affect parallel fluid flow. In
particular, fluid flow within an array of parallel channels is
controlled by a single pair of electrodes. As shown, substrate 302
includes a plurality of parallel channels 304-332. These parallel
channels each terminate in transverse channels 334 and 336.
Transverse channel 334 has a tapered width, going from its widest
at the point where it intersects the nearest parallel channel 304
to the narrowest at the point it intersects the most distant
parallel channel 332. Transverse channel 336, on the other hand,
goes from its widest at the point it intersects parallel channel
332, to the narrowest where it intersects parallel channel 302.
Electrodes are included in the ports 338 and 340 at the wider
termini of transverse channels 334 and 336, respectively. The
dimensions of these tapered channels are such that the current flow
delivered through each of the parallel channels, via the tapered
channels, is substantially equal, thereby permitting equal flow
rates in each of the parallel channels. As shown, transverse or
sample introduction channel 342 is oriented so that it crosses each
parallel channel at the same point relative to one or the other
electrode, to ensure that the potential at the intersection of
transverse channel 342 is the same in each of the parallel
channels, again, to prevent the formation of transverse electrical
fields, or "shorting out" the array of channels. This results in
the sample introduction channel 342 being disposed across the
parallel channels at a non-perpendicular angle, as shown.
[0060] In operation, a sample fluid, e.g., disposed in port 344, is
flowed through transverse channel 342, and across the intersection
of the parallel channels 304-332 by applying a potential across
ports 344 and 346. Once the sample is disposed across the one or
more desired parallel channels, e.g., as dictated by the serial to
parallel conversion desired (see FIGS. 2A-2D), a potential is then
applied across ports 338 and 340, resulting in an equal fluid flow
through each of the parallel channels and injection of the sample
fluid into each of the desired parallel channels.
[0061] FIG. 5 illustrates still another embodiment for practicing
the principles set forth herein. In this embodiment, a substrate
includes a large number of parallel channels. For ease of
discussion, these channels are referred to herein as parallel
channels 404-410, although it should be understood that preferred
aspects will include upwards of 50, 100, 500 or more separate
parallel channels. The parallel channels 404-410 terminate at one
end in transverse channel 412 and at the other end in transverse
channel 414. Electrodes are provided within ports 416 and 418, and
420 and 422 at the termini of these transverse channels. In this
embodiment, the problems of varying current within the different
parallel channels are addressed by providing transverse channels
412 and 414 with sufficient width that current variation across the
length of these transverse channels, and thus, as applied to each
parallel channel, is negligible, or nonexistent. Alternatively, or
additionally, a single electrode may be disposed along the length
of each of these transverse channels to ensure equal current flow
at the transverse channel's intersection with each parallel
channel.
[0062] As shown, however, transverse or sample introduction channel
424 intersects each of the parallel channels, and is controlled by
electrodes disposed within ports 426 and 428 at the termini of
channel 424. As described for FIG. 4, above, the sample
introduction channel intersects each parallel channel at a point
where the potential applied to each channel will be equal. In this
aspect, however, the channel is arranged substantially parallel to
transverse channels 412 and 414, as each parallel channel is
subjected to the same current.
[0063] In operation, a sample, e.g., disposed in port 426, is
flowed through sample channel 424, across the intersection of the
various parallel channels 404-410, by applying a potential across
ports 426 and 428. Once the sample fluid is in its appropriate
location, i.e., across all or a select number of parallel channels,
a potential is applied across ports 416:420 and 418:422, injecting
a plug of sample into the parallel channels.
[0064] An alternate, although similar channel geometry for a serial
to parallel conversion device is shown in FIG. 8. As shown the
device includes an array of parallel channels where each channel is
coupled between two reservoirs/electrical nodes (CBR and CBW).
Equal currents are applied to the parallel channels via fractal
channel networks at each end of the parallel channel array. Sample
is introoduced via a central channel introduction channel (disposed
between and coupled to reservoirs SBW and SBR). Additional channels
are provided to introduce additional elements to the sample
material, or additional samples, prior to introduction into the
parallel array (channels disposed between and coupled to reservoirs
SR-1 and SW-1 or SR-2 and SW-2). The fractal channel networks have
wider cross-sections in order to ensure optimal current is
delivered to the parallel channel array, e.g., minimize
resistance.
[0065] Although the present invention is exemplified in terms of
utilizing either a parabolic channel geometry or a wide channel
geometry to ensure equal resistances in parallel channels, it will
be readily appreciated that both accomplish substantially the same
goals, and further that a combination of wider channel geometries
and parabolic channel geometries in those channels connecting the
termini of the parallel channels, is often used to optimize for
maintaining applied currents in an acceptable range (wider channels
require greater applied currents) and minimizing the use of
substrate area (parabolic channel geometries typically utilize
greater substrate areas than straighter channels). Thus, in
preferred aspects,
EXAMPLES
[0066] 1. Parallel Transport of Fluorescent Material in
Interconnected Parallel Channels
[0067] A microfluidic device was fabricated from a glass base
substrate having another glass substrate overlaying the first. The
device included a channel layout as shown in FIG. 4. The device was
filled by capillary action using a sodium tetraborate buffer placed
into one of the reservoirs. Fluorescein mixed with the running
buffer, was placed into reservoir 344 and drawn across the
intersections with the parallel channels by applying a voltage
gradient between reservoirs 344 and 346.
[0068] Once the fluorescein filled the entire transverse channel
342, the voltage gradient was changed from between reservoirs 344
to 346, to between reservoirs 338 and 340. Because of the geometry
of the transverse channels 334 and 336, as well as the angled
geometry of transverse channel 342, the amount of current passing
through each of parallel channels 304-332 was maintained
substantially equal, and the fluorescein plugs were transported
down their respective channels at substantially the same rate.
[0069] FIG. 6 is a photograph illustrating the introduction of
fluorescein into the array of parallel channels. As can be seen
from this photograph, the fluorescent plugs in each channel are
moving at substantially the same rate.
[0070] 2. Parallel Analysis of Nucleic Acid Fragments in a
Microfluidic System
[0071] A microfluidic device fabricated from two bonded glass
layers, and having the channel geometry shown in FIG. 8, was used
to analyze the same mixture of nucleic acid fragments in 32
parallel channels, where the samples were serially introduced into
the device via the central sample channel. Currents were maintained
substantially constant from one parallel channel to the next, by
using only two reservoir/electrodes to control material movement in
those parallel channels. The two reservoir/electrodes (labeled CBR
ad CBW) were connected to the parallel channels via corresponding
fractal channel networks. The sample to be tested was placed in the
sample well (SR-2) and transported across the parallel channel
array by transporting the sample to the sample waste well
(SBW).
[0072] The sample solution was a combination of 3.5% Genescan
polymer sieving matrix in the Genescan buffer, a 500:1 dilution of
the SYBR Green intercalating dye, and 5:1 dilution of a 1
.mu.g/.mu.L solution of double-stranded DNA size standard,
.PHI.X174 cleaved with HaeIII (Promega Corp.). The microfluidic
device was first filled with a 3.5% solution of the GeneScan.TM.
polymer in the Genescan buffer, including a 500:1 dilution of the
SYBR Green intercalating dye. This solution was placed in all
wells, except well SR-2, which was filled with the DNA sample
solution.
[0073] The experiment consisted of first filling the loading
channel with DNA solution by applying a potential between wells
SR-2 and SBW. A small amount of sample was then injected in the
parallel separation channels by briefly applying a potential
between wells CBW and CBR. The loading channel was then cleared out
by applying a potential between wells SW-2 and SBW. This clearing
out ensured that only a limited amount of DNA was injected into
each separation channel. Finally, the separation was done in each
channel simultaneously, by again applying an electric potential
between wells CBW and CBR. The separation was observed at a
location at the bottom of the separation channels, near well
CBW.
[0074] Detection of separated species was carried out using a
single detector utilizing a Nikon inverted Microscope Diaphot 200,
with a PTI Model 814 PMT detection system, for epifluorescent
detection. An Opti-Quip 1200-1500 50W tungsten/halogen lamp coupled
through a 40.times. microscope objective provided the light source.
Excitation and emission wavelengths were selected with a FITC
filter cube (Chroma, Brattleboro Vt.) fitted with appropriate
filters/dichroic mirrors. Five separate runs were performed where
sample was loaded through the main channel and simultaneously
injected into each of the 32 parallel separation channels. In each
separate run, detection was carried out at a different one of the
32 channels.
[0075] The separation of the different fragments in each of five
different parallel channels is shown in FIG. 7, as retention time
vs. fluorescent intensity. The retention time is shown as the time
from the first injection. The specific channel is indicated for
each separation. Since only a single detector was used, each plot
is a different sample of one of the 32 separations that were in
fact done simultaneously each time a sample was injected. The
similarity of each scan indicates that similar amounts of material
were injected into each channel, and further, that relative
retention times, e.g., electrophoretic mobilities, in each channel
were maintained substantially constant.
[0076] While the foregoing invention has been described in some
detail for purposes of clarity and understanding, it will be clear
to one skilled in the art from a reading of this disclosure that
various changes in form and detail can be made without departing
from the true scope of the invention. All publications and patent
documents cited in this application are incorporated by reference
in their entirety for all purposes to the same extent as if each
individual publication or patent document were so individually
denoted.
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