U.S. patent application number 11/595270 was filed with the patent office on 2007-03-08 for microfluidic device with controlled substrate conductivity.
This patent application is currently assigned to Caliper Life Sciences, Inc.. Invention is credited to Luc J. Bousse, Richard J. McReynolds, Seth R. Stern.
Application Number | 20070053799 11/595270 |
Document ID | / |
Family ID | 37886038 |
Filed Date | 2007-03-08 |
United States Patent
Application |
20070053799 |
Kind Code |
A1 |
Bousse; Luc J. ; et
al. |
March 8, 2007 |
Microfluidic device with controlled substrate conductivity
Abstract
A method to achieve controlled conductivity in microfluidic
devices, and a device formed thereby. The method comprises forming
a microchannel or a well in an insulating material, and ion
implanting at least one region of the insulating material at or
adjacent the microchannel or well to increase conductivity of the
region.
Inventors: |
Bousse; Luc J.; (Los Altos,
CA) ; Stern; Seth R.; (Mountain View, CA) ;
McReynolds; Richard J.; (San Jose, CA) |
Correspondence
Address: |
CALIPER LIFE SCIENCES, INC.
605 FAIRCHILD DRIVE
MOUNTAIN VIEW
CA
94043-2234
US
|
Assignee: |
Caliper Life Sciences, Inc.
Mountain View
CA
|
Family ID: |
37886038 |
Appl. No.: |
11/595270 |
Filed: |
November 10, 2006 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
|
|
10384349 |
Mar 7, 2003 |
|
|
|
11595270 |
Nov 10, 2006 |
|
|
|
60362340 |
Mar 8, 2002 |
|
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Current U.S.
Class: |
422/400 |
Current CPC
Class: |
B81B 7/0006 20130101;
G01N 27/44791 20130101; B81C 1/00698 20130101; Y10T 436/11
20150115; B01L 2400/0415 20130101; Y10T 436/2575 20150115; B01L
2300/0645 20130101; G01N 27/44743 20130101; B01L 2200/12 20130101;
Y10T 29/49002 20150115; Y10T 29/49 20150115; B81B 2201/058
20130101; B81C 2201/019 20130101; B81B 2203/0338 20130101; B01L
3/502707 20130101 |
Class at
Publication: |
422/100 |
International
Class: |
B01L 3/02 20060101
B01L003/02 |
Claims
1. A microfluidic device, comprising: a first layer made of an
insulating material; a first microchannel formed in said first
layer; at least one of a second microchannel and a well formed in
said first layer; and at least one ion implanted region in said
first layer located at or adjacent the first microchannel and said
at least one of said second microchannel and said well, said at
least one ion implanted region providing a conductive path between
said first microchannel and said at least one of the second
microchannel and the well.
2. The device of claim 1, wherein said insulating material
comprises a silica-based material.
3. The device of claim 1, wherein said insulating material
comprises a polymer material.
4. The device of claim 1, wherein said first layer comprises a
substrate of the microfluidic device.
5. The device of claim 1, further comprising a second layer bonded
to said first layer.
6. The device of claim 1, wherein said first layer comprises a
cover plate.
7. The device of claim 1, wherein said insulating material
comprises a silica-based material.
Description
CROSS REFERENCE TO RELATED APPLICATIONS
[0001] This is a continuation of U.S. application Ser. No.
10/384,349, filed Mar. 7, 2003, which claims the benefit of U.S.
Provisional Application Ser. No. 60/362,340, filed Mar. 8, 2002,
which is incorporated herein by reference in its entirety for all
purposes.
BACKGROUND OF THE INVENTION
[0002] 1. Field of the Invention
[0003] The present invention is related to microfluidic devices,
and specifically to methods for modifying the conductivity of
materials used in the fabrication of those devices.
[0004] 2. Related Art
[0005] Microfluidic technology enables the miniaturization and
automation of many laboratory processes. Devices employing
microfluidic technology can integrate the power of an entire
laboratory full of equipment and people into a single
"lab-on-a-chip." Each microfluidic device (hereafter also referred
to as a "chip") contains a network of microscopic channels, or
microchannels, through which fluids can be moved and in which
experiments can be performed. The design of microfluidic devices
for biochemical applications involves the disciplines of fluid
dynamics, biochemistry, software, and thin film manufacturing.
[0006] In microfluidic devices, the driving forces that move fluids
within the channels of the device can be electrokinetic forces,
pressure forces, or a combination of the two. Electrokinetic forces
are typically generated by applying an electric field across a
microchannel, where the direction of the field is parallel to the
desired direction of fluid flow. The electric field is typically
applied by placing electrodes in reservoirs at the ends of the
microchannel, and applying a voltage across the electrodes with a
computer-controlled power supply. The voltage applied across the
electrodes produces fluid flow via one or both of the phenomena of
electroosmosis or electrophoresis. Electroosmosis occurs when an
electric field is applied across a channel whose surface or walls
contain charged functional groups. The charge on the channel wall
ionizes a thin layer of fluid near the wall. This thin layer of
ionized fluid is attracted to one of the electrodes, creating a
flow of ionized species toward that electrode. The flow of ionized
species produces both a bulk fluid flow and an electrical current.
The bulk flow rate through a microchannel can be controlled with a
high degree of precision by controlling the electrical current that
accompanies the flow through the microchannel. The other phenomena
that produces electrokinetic flow, electrophoresis, is the movement
of charged molecules or particles in a fluid subjected to an
electric field. Electrophoresis can be used to move charged
molecules in solution, or to separate charged molecules that have
different electrophoretic mobilities (which is roughly their charge
to mass ratio). Electrophoresis and electroosmosis often occur at
the same time when an electric field is applied to a microchannel.
Techniques have been developed for minimizing one electrokinetic
force while maintaining the other, as appropriate, for a given
application. Precise control over fluid flow within microchannels
requires precise control of the driving forces, such as
electrokinetic or pressure forces. Precise control over fluid flow
also requires precise engineering of the microchannels themselves
because fluid flow also depends on channel geometry and surface
properties.
[0007] Microfluidic devices are typically fabricated by etching or
embossing grooves into a substrate, and then affixing a cover to
the substrate to form the microchannels. In most microfluidic
devices that employ electrokinetic flow, both the substrate and the
cover plate are made of an insulating material such as glass.
Insulating materials help reduce the electrical current leakage
between microchannels. By reducing current leakage between
microchannels, the use of insulating materials allows an increased
packing density of components, such as microchannels, in a
microfluidic device.
[0008] In some applications, it may be advantageous to allow a
localized leakage of current between different channels in a
microfluidic device. The leakage of current between channels allows
the electrical currents that drive electrokinetic flow to flow in
directions other than parallel to the length of the microchannels.
In other words, having a conductive path between channels provides
the ability of initiating electrokinetic flow in directions other
than along the length of a channel. For example, fluid could be
made to flow into the sidewall of a channel. Microfluidic devices
with a conductive path between channels could provide advantages
over standard microfluidic devices in the areas of sample
concentration and two-dimensional separation.
[0009] One set of researchers has fabricated microfluidic devices
that employ electrical current leakage between microchannels for
the purpose of concentrating samples. Khandurina, J., et al., Anal.
Chem. 71, pp. 1815-1819 (1999). In these microfluidic devices, the
current leaks between microchannels through a porous membrane. The
porous membrane is a separate layer of material sandwiched between
the cover plate and substrate of a microfluidic device. In the
devices shown in Khandurina, fluid from a main channel that
terminates at a "T" shaped intersection with a separation channel.
The fluid from the main channel is made to flow straight into the
opposing wall of the "T" shaped intersection by allowing electrical
current to flow into the opposing wall through a porous membrane
above the wall. By flowing sample from the main channel into the
opposing wall, the sample accumulates, and thus concentrates, at
the "T" intersection. When enough sample has accumulated at the
intersection, the sample is directed to flow down the separation
channel. The device in Khandurina could be useful in assays in
which a sample to be separated into components must be concentrated
in order to increase the concentration of at least some of the
components above a detectable threshold.
[0010] There are several problems with microfluidic devices that
employ porous membranes to provide conductive paths between
microchannels. First, the lifetime of these devices is short and
unpredictable due to the nature of the porous membrane. Second, the
resistance of the porous membranes may change with time. Third, the
process for fabricating porous membranes lacks the dimensional
control needed to fabricate porous membranes between closely spaced
microchannels. Fourth, the nature of the conductivity of the porous
membrane is not certain, and that could lead to unexpected
fluctuations of conductivity both between and within microfluidic
devices. Finally, having a conductive path between microchannels
may prevent the manufacture of devices with densely packed
microchannels.
[0011] Given the limitations of porous membranes, it is desirable
to have an alternative method of providing conductive paths between
microchannels in a microfluidic device. It would be particularly
desirable if the conductive paths could be provided in a way that
does not require the addition of an extra layer of material, such
as the above-described layer of a porous membrane material, to the
microfluidic device structure. Furthermore, it would be desirable
that the dimensions of the conductive paths be able to be precisely
and accurately defined. It would also be desirable that the degree
of conductivity between channels be controllable. In its various
aspects, embodiments of the present invention provide these and
other advantages over currently known methods of allowing current
to flow between the channels of a microfluidic device.
BRIEF SUMMARY OF THE INVENTION
[0012] The present invention is directed to a microfluidic device
comprising at least two microchannels formed from grooves in an
insulating substrate, and at least one ion implanted region in the
insulating substrate located between the grooves forming the
microchannels, the at least one ion implanted region having
increased conductivity compared to the insulating substrate.
[0013] The present invention is also directed to a method to
achieve controlled substrate conductivity in microfluidic devices,
and devices formed thereby. The method comprises forming a
microchannel in an insulating substrate, and ion implanting at
least one region of the insulating substrate at or adjacent the
microchannel to increase conductivity of the region. In some
embodiments of the invention, the insulating substrate is a
silica-based material, whereby the ion-implanting step increases
the conductivity of the silica-based material in at least one
region. In alternative embodiments, the insulating substrate is a
polymer material, whereby the ion-implanting step increases the
conductivity of the polymer material in at least one region.
[0014] By providing regions where substrate conductivity is
increased, it is possible to run loading currents through the
substrate, and thus accumulate sample components. The
ion-implantation process used in embodiments of the invention can
be accurately and precisely modify the conductivity of small areas
of an insulating substrate, so that the invention is compatible may
be employed on a microfluidic device with closely packed
microchannels.
[0015] These and other advantages and features will become readily
apparent in view of the following detailed description of the
invention.
BRIEF DESCRIPTION OF THE DRAWINGS
[0016] The features and advantages of the present invention will
become more apparent from the detailed description set forth below
when taken in conjunction with the drawings in which like reference
numbers indicate identical or functionally similar elements.
Additionally, the left-most digit of a reference number identifies
the drawing in which the reference number first appears.
[0017] FIG. 1 is an exploded view of a microfluidic device in
accordance with the invention.
[0018] FIGS. 2 and 3 illustrate methods of forming regions of
increased conductivity in microfluidic chips in accordance with the
present invention.
[0019] FIG. 4 illustrates a microfluidic device in accordance with
the present invention.
[0020] FIG. 5 is a magnified view of a portion of the microfluidic
device of FIG. 4.
DETAILED DESCRIPTION
[0021] Embodiments of the present invention will now be discussed
in detail. While specific features, configurations and arrangements
are discussed, it should be understood that this is done for
illustration purposes only. A person skilled in the relevant art
will recognize that other steps, configurations and arrangements
may be used without departing from the spirit and scope of the
invention. It should be appreciated that the microfluidic devices
in accordance with the present invention can be used to perform a
variety of experiments and operations, and thus the techniques
described herein could be used in connection with devices for such
other functions.
[0022] An exploded view of a microfluidic device in accordance with
the invention is shown in FIG. 1. The microfluidic device 10
comprises two layers: a substrate 12 and a cover plate 18. The
substrate 12 may be made of a variety of materials, including
quartz, glass, polymer, ceramic or even semiconductor materials.
The substrate 12 comprises a pattern of grooves 16 on the upper
surface 14 of the substrate. The pattern of grooves 16, when
enclosed by the cover plate 18, will form the pattern of
microfluidic channels in the assembled microfluidic device. The
pattern of grooves may be formed by a variety of manufacturing
methods, many of which are used in the semiconductor industry. For
example, the pattern of grooves may be formed by embossing the
pattern onto a polymer substrate, injection molding a polymer into
a substrate containing the pattern, or by a combination of
lithography and etching. In one embodiment, the pattern of grooves
16 is defined using a lithography process, and etched into the
substrate using a wet etch process. A process combining lithography
and wet etch is able to create highly precise microchannels with
dimensions that can be varied by width and depth. A typical
microchannel is roughly 50 .mu.m wide and 10 .mu.m deep.
[0023] After the pattern of grooves 16 is formed on the top surface
14 of the substrate 12, a cover plate 18 is fused with the
substrate 12. Such fusion can be performed using a variety of known
bonding techniques, including thermal and anodic bonding. The cover
plate 18 may be formed from a variety of materials, including
quartz, glass, polymer, ceramic or semiconductor materials. The
cover plate 18 encloses the pattern of grooves 16 formed in the
substrate 12 and converts them to microfluidic channels, or
microchannels. Either or both of the substrate 12 or cover plate 18
may include holes or apertures disposed therein. In the embodiment
shown in FIG. 1, holes 24 in the cover plate 18 form reservoirs or
wells that are disposed above and fluidly connected to the
unintersected termini of the grooves in the substrate 12. Fluids
may be introduced into the microchannels of the assembled
microfluidic device through these reservoirs. The size of an
assembled microfluidic device can vary from less than one inch to a
few inches on a side. The assembled devices are typically packaged
in plastic holders, which make them easier for the user to
handle.
[0024] Other embodiments of microfluidic devices are compatible
with the present invention. For example, instead of only having
holes in the cover layer, microfluidic devices in accordance with
the invention may have holes disposed through the substrate or
through both the substrate and cover plate. The extra holes could
either provide separate reservoirs on opposing sides of the
microfluidic device or to provide through-holes that provide fluid
access to the channels of the device. Embodiments of microfluidic
devices employing holes in the substrate or both the substrate and
cover layer are in U.S. Pat. Nos. 5,779,868 and 6,090,251, both of
which are assigned to the assignee of the present invention. Other
embodiments of microfluidic devices that are compatible with the
present invention include multilayer microfluidic devices, which
comprise two or more substrate layers. The microchannel formed in
the various layers of a multiplayer microfluidic device can be
interconnected using vias or through-holes.
[0025] Microfluidic devices, or chips, are currently commercially
available in two basic formats: planar and sipper. In planar chips,
such as the chip shown in FIG. 1, the user introduces all chemical
reagents, including samples, into reservoirs on the chip. Planar
chips are sold for use with the Agilent 2100 Bioanalyzer system.
These chips include the LabChip.RTM. DNA Analysis, RNA Analysis,
Protein Analysis and Cell Fluorescence Analysis chips, which are
manufactured by Caliper Technologies Corp., of Mountain View,
Calif. Because samples are deposited into separate reservoirs in
planar chips, the number of samples that can be analyzed by a
planar chip is typically limited by the number of sample reservoirs
provided on that chip. While planar chips could be washed and
potentially reused, they are generally discarded after use.
[0026] Sipper chips are designed to analyze a large number of
samples, which makes sipper chips useful for high throughput
applications. In the sipper chips, minute quantities of a large
number of samples can be tested in a single chip. The samples are
introduced into the capillary one after the other, spaced by buffer
solution. The samples then proceed through the microchannel network
in a continuous flow, assembly-line fashion. A typical sipper chip
employs one or more integrated sample accession capillaries for
interfacing with an external collection of sample sources, such as
a multiwell plate. The sample accession capillary is typically a
small glass tube inserted into the substrate portion of the chip.
Embodiments of sipper chips compatible with the present invention
are described in U.S. Pat. No. 5,779,868, which is assigned to the
assignee of the present invention. Commercially available sipper
chips include chips used by the Caliper AMS 90 and 250 HTS
systems.
[0027] In most microfluidic devices, fluids are moved through the
microchannels of the device by means of electrokinetic forces,
pressure forces, or a combination of the two. Electrokinetic forces
are typically generated by applying an electric field along the
length of a microchannel, parallel to the desired direction of
fluid flow. For example, in the microfluidic device 10 in FIG. 1,
electrokinetic driving forces would be applied to the
microchannels, which are formed by enclosing the grooves 16, by
placing electrodes in the reservoirs 24 and applying voltages
between the various electrodes. Fundamental techniques for
controlling electrokinetic flow in the microchannels of a
microfluidic device were invented by Dr. J. Michael Ramsey. These
techniques are covered by a series of issued and pending patents,
including U.S. Pat. Nos. 6,001,299 and 5,858,195. Dr. Ramsey's
techniques control fluid flow within multiple microchannels can by
simultaneously applying separately controllable electric fields
across the different microchannels. Software programs can be
written for computer controlled power supplies to generate highly
specific and complex networks of flow within a network of
microchannels.
[0028] This invention is directed to using ion-implantation to
controllably and locally increase the conductivity of a substrate
or cover plate of a microfluidic device. By increasing the
conductivity in a defined area, a path for electrical current can
be defined between microchannels. The physical separation distance
between microchannels in a microfluidic device in accordance with
the invention can range from between 10-100 .mu.m, and in most
embodiments between 20 to 50 .mu.m. The ion-implantation process
provides control over the conductivity of the region of the
microfluidic device being implanted by controlling the dose,
energy, and subsequent thermal annealing of that region.
Ion-implantation enables localized high-conductivity areas to be
formed on a chip without degrading performance elsewhere on the
chip.
[0029] As will be described below, ion implanting according to the
present invention can be performed in various areas or regions of
the substrate and/or cover plates of a microfluidic device. When
referencing the ion implanting of an area or region, the terms
"adjacent a microchannel" and "adjacent a reservoir", or the like
phrases, are used herein to mean a variety of possible relative
positioning of the ion implanted area or region and a microchannel
and/or reservoir. As examples, "adjacent" can mean that the
ion-implanted region: fully or partially overlaps (i.e., is fully
or partially integral) with a portion of a microchannel and/or
reservoir; or is separated a distance from a portion of a
microchannel and/or reservoir. Such a separation distance or
overlap will be implementation dependent.
[0030] Various authors have described methods to change the
conductivity of glass (or of fused silica) by ion implantation (see
Okura, T., and Yamashita, K, Solid State Ionics 136 SI:1049-1054
(2000); Rebohle, L., et al., Applied Physics B-lasers and Optics
71:131-151 (2000); Nakajima, A., et al., J. Vacuum Sci. Technol. B
17:1317-1322 (1999); Hosono, H., et al., J. Non-crystalline Solids
182:109-118 (1995); Prawer. S., et al., J. Appl. Physics
73:3841-3845 (1993); and Martin, P., et al., J. Appl. Physics
72:2907-2911 (1992)). Examples of implanted species that have been
used are: protons, sodium, antimony, silicon, germanium, carbon,
titanium, and chromium. Ion implantation allows accurate control of
dose and depth, and has long been a vital part of silicon
integrated circuit technology, where such control is essential. The
details of ion implantation techniques, including thermal annealing
and the manufacturing equipment to carry out ion implantation would
be apparent to a person skilled in the relevant art.
[0031] To localize the effect of an ion-implantation process, some
form of masking is typically used. This masking can be accomplished
using lithography techniques that employ either positive or
negative photoresist materials. Such lithography techniques are
commonly used in the integrated circuit industry, and have been
applied in the manufacture of flat panel displays, circuit boards,
microfluidic devices, and various integrated circuits. When a
lithography process is used to pattern a substrate, the substrate
is first coated with one or more layers of a photoresist material.
In some embodiments, the substrate may be coated with a layer of
chrome before the photoresist is applied. The chrome may act as an
adhesion layer between the photoresist and substrate materials to
which the photoresist does not adequately adhere. The substrate is
then placed in an aligner, in which the substrate is placed on a
stage and held in place by a chuck. The chuck is typically a vacuum
or electrostatic chuck capable of securely holding the substrate in
place. The photoresist on the substrate is exposed to an image
projected onto its surface by passing radiation through a patterned
mask or reticle. As is known to those skilled in the relevant art,
the radiation could be visible light, UV light, x-rays, ions, or
electrons.
[0032] The projected image produces changes in the characteristics
of the coating of photoresist material. These changes occur in the
portions of the photoresist that were exposed to radiation during
exposure. Subsequent to exposure, the layer is developed to produce
a patterned layer of photoresist. In some embodiments, the
substrate covered with the patterned layer of photoresist is then
subjected to an etching process. Some areas of the photoresist
pattern expose the underlying substrate from the etching process,
while other portions of the pattern shield the substrate from the
etching process. Accordingly, the photoresist pattern is
effectively transferred to the underlying substrate. As previously
discussed, this combination of lithography and etching can be used
to form grooves in a glass or polymeric substrate that, when
covered, become the microchannels of a microfluidic device. In
another aspect of the invention, a photoresist pattern formed by a
lithography process is used to shield portions of a substrate or
cover plate from an ion implantation process.
[0033] As the size of a substrate increases, the equipment required
to pattern the entire substrate at once becomes more expensive. So
rather than expose the entire substrate at one time, sub portions
of the photoresist layer are exposed one at a time. A special type
of aligner known as a "step-and-scan" aligner is designed to expose
only a portion of a substrate at a time. A step-and-scan aligner
contains a projection optics system that has a narrow imaging slot.
An entire substrate can be exposed by placing the imaging slot and
reticle over different portions of the substrate. To accomplish
this, the stage on which the wafer sits is then moved between
exposures to allow multiple copies of the reticle pattern to be
exposed over the substrate surface. In this manner, the sharpness
of the image projected onto the substrate is maximized. Using a
step-and-scan technique generally assists in improving overall
image sharpness. For more background see, Nonogaki et al.,
"Microlithography Fundamentals in Semiconductor Devices and
Fabrication Technology" (Marcel Dekker, Inc.: New York, N.Y. 1998).
Step-and-repeat and field stitching lithography techniques can also
be used.
[0034] An exemplary method for modifying the conductivity of a
portion of a microfluidic device is shown in FIG. 2. This method
employs the lithography and ion implantation processes described
above. For clarity, the method in FIG. 2 will be described in terms
of its application to the glass substrate portion of a microfluidic
device. One skilled in the relevant art would recognize that
methods in accordance with the invention could be applied to other
portions of a microfluidic device, such as a cover plate, and could
be applied to substrates and cover plates made of materials other
than glass.
[0035] The first step in the method of FIG. 2 is the fabrication of
the pattern of grooves in the substrate that will, when covered,
form the microfluidic channels in a microfluidic device. The
pattern of grooves is formed in step 202 by means of the
combination of lithography and etching described above. The
substrate is made of an insulating material such as glass, a
silica-based material or a polymeric-based material. Before
proceeding to the next step, any residual layers, such as chrome
and photoresist, are removed using known techniques.
[0036] In a step 204, a thick photoresist such as Shipley SPR 220
or Clariant AZ EXPLOF 5000 is applied and patterned by exposing and
developing the photoresist. In some embodiments it may be
advantageous to soft bake the photoresist after it is applied to
the substrate. The portions of this layer of photoresist that are
not removed during the developing process will shield the portions
of the substrate they cover from the ion implantation process in a
step 206. In some embodiments it may be advantageous to hard bake
the photoresist after it is developed. During ion implantation 206,
the exposed portions of the substrate, i.e. the portions of the
substrate not covered by photoresist, will have their conductivity
increased by the implantation of ions. The portions of the
substrate with increased conductivity regions are also referred to
herein as "glass resistors." The degree to which the conductivity
of the glass resistors increases during ion implantation depends on
the dose and energy of the implanted ions. After ion implantation
is complete, the photoresist layer is stripped away 208. Due to
possible hardening of the photoresist during ion implantation,
especially during after high dose implants, stripping the
photoresist 208 may require plasma treatments in addition to the
standard wet chemical baths. As would also be apparent to a person
skilled in the relevant art, it may be desirable to thermally
anneal the substrate to repair damage to the substrate and to
electrically activate the implanted ions. Finally, at a step 210
the substrate, which now comprises a pattern of grooves and one or
more glass resistors, is bonded to a cover plate.
[0037] To completely avoid the problems associated with hardening
of photoresist during ion implantation, a material other than
photoresist may be used to shield portions of the substrate from
the ion implantation process. In other words, a layer of material
could be deposited on the substrate to form a masking layer. The
material forming the masking layer could be a metal, or an
insulator material different than the substrate material. Specific
examples of materials suitable for use as a masking layer are
chrome, silicon nitride, amorphous silicon and polysilicon. The
masking layer would form a pattern that covers the portions of the
substrate that are to be shielded from the ion-implantation
process, and leaves exposed the portions of the substrate that are
to be implanted with ions. The masking layer is typically patterned
by means of a lithography process. In an exemplary embodiment, a
layer of photoresist is patterned (i.e. applied, developed, and
exposed) so that only the portions of the substrate that are to be
covered by the masking layer are left exposed. Next, a layer of
chrome is sputtered onto the substrate. Finally, the resist is
stripped so that only the chrome deposited on the exposed areas of
the substrate remains. The process used to deposit chrome in this
exemplary embodiment is commonly known as a lift-off process. After
the ion implant is carried out, the masking layer can be removed
using a selective etch process so as not to affect the underlying
substrate.
[0038] A second exemplary method for modifying the conductivity of
a portion of a microfluidic device is shown in FIG. 3. In the
embodiment of FIG. 3 the substrate is subjected to ion-implantation
before the pattern of grooves is formed on the substrate surface.
In a step 302 shallow alignment marks are etched at the edges of a
substrate. These alignment marks facilitate the proper alignment of
the ion-implanted regions, the glass resistors, with the yet-to-be
formed pattern of grooves. In a step 304, a layer of photoresist is
applied to the substrate, and is then exposed and developed. This
layer of photoresist, just like the layer of photoresist in step
204 of FIG. 2, serves to shield portions of the substrate from the
ion implantation process. As was discussed with regards to the
layer of photoresist in FIG. 2, the photoresist in step 304 of FIG.
3 may harden during the ion implantation process. Accordingly, in
some embodiments it may be desirable to replace the layer of
photoresist in step 304 with a masking layer. In a step 306, ion
implantation is performed to form one or more glass resistors. In a
step 308 the photoresist is stripped. As discussed with regards to
the embodiment in FIG. 2, it may be desirable to thermally anneal
the substrate after implantation. In a step 310 a second layer
photoresist is applied to substrate, and is then exposed and
developed. This layer of photoresist defines the pattern of grooves
that will form the microchannels in the finished microfluidic
device. In a step 312, the pattern of grooves is etched into the
surface of the substrate. In a final step 314, the photoresist is
stripped from the substrate. The substrate and cover plate can then
be bonded together to form a microfluidic device.
[0039] In other embodiments of the invention, the conductivity of
portions of the cover plate are modified by subjecting the cover
plate to an ion implantation process such as one of those shown in
FIGS. 2 and 3. In still other embodiments, the conductivity of
portions of both the cover plate and the substrate are
modified.
[0040] Glass resistors formed by ion implantation according to the
present invention can be used is a variety of ways in microfluidic
devices. One exemplary benefit that can be achieved by employing
glass resistors in a microfluidic device is increased sensitivity
in a protein assay. Glass resistors in accordance with the present
invention can employed in a protein assay chip in the Agilent 2100
Bioanalyzer, for example.
[0041] FIG. 4 illustrates a 16-well protein assay microfluidic chip
400 in accordance with the present invention. The wells or
reservoirs 406, 408, 416-436, 440, 450, 452, 484 are in fluid
communication with the network of microchannels, which includes
microchannels 460-470. Of the sixteen wells, ten 416-436 contain
samples to be analyzed, two 408, 484 are waste wells, one 440
supplies a reagent such as a fluorescent dye that enables detection
of selected species, and one 406 is a source of buffer. These
fourteen wells are fluidly connected by a first network of
channels. The remaining two wells 450,452 are fluidly connected by
a second network of channels. In this embodiment, the first and
second networks of channels are not fluidly connected. A region of
increased conductivity, a glass resistor, is shown generally at a
region 490. FIG. 5 is a magnified view of the region adjacent the
glass resistor 490, which is enclosed by a box for illustrative
purposes.
[0042] The basic function of chip 400 is to separate a sample into
its components by electrophoretic means. Means of electrophoretic
separation that may be employed in embodiments of the invention are
described in U.S. Pat. No. 5,948,227, which is assigned to the
assignee of the present invention. In the device of FIGS. 4 and 5,
the electrophoretic separation takes place in separation channel
404. Samples from wells 416-424 are injected into separation
channel 404 via channels 462 and 472, while samples from wells
428-436 are injected into separation channel 404 via channels 464
and 472. The electrophoretic separation of the sample takes place
as the sample travels through the separation channel 404 from the
intersection of channels 472 and 404 to waste well 408. In this
exemplary embodiment, only one sample at a time is separated in the
separation channel 404. To improve device throughput, however, a
sample can be preloaded into channel 472 while a previously
injected sample is being separated in separation channel 404. The
preloading process does not interfere with the separation taking
place in separation channel 404 because the flow of the sample
being preloaded is diverted into channel 460, which empties into
waste well 484. Preloading a subsequent sample in this manner
minimizes the time required to load the sample into the separation
channel 404. This type of preloading is described in more detail in
U.S. Pat. No. 5,948,227, and is implemented in DNA and RNA assay
chips for the Agilent 2100 Bioanalyzer.
[0043] To inject a sample into the separation channel 404, the
sample travels from its well through either channel 462 or 464 into
channel 472. The sample is propelled through those channels by
electrokinetic forces generated by voltages applied between
electrodes (not shown) immersed in the sixteen reservoirs of the
microfluidic device 400. When the sample arrives at the "T"
intersection between channels 472 and 404, the sample is directed
to travel straight out of channel 472, across channel 404, into the
opposite wall of channel 404. The electrical current directing the
flow of sample into the wall flows through channel 472, through the
glass resistor 490, and finally through channel 468. The glass
resistor allows electrical current to flow from channel 472 into
channel 468, even though channels 472 and 468 are not in fluid
communication. The desired electrical currents are supplied via
power-supply electrodes immersed in the sixteen reservoirs of the
microfluidic device 400. If, for example, the sample being injected
into separation channel 404 originated from reservoir 418, the
desired electric current would be generated by applying appropriate
voltages to electrodes in reservoirs 418, 450, and 452. Note that
voltages may have to be simultaneously applied to other reservoirs
connected to the first channel network to prevent the flow of other
samples into channels 462, 464 and 472, and to prevent the
diversion of sample into waste well 484. Also, a voltage may have
to be applied across the length of separation channel 404, by means
of voltages applied to electrodes in reservoirs 406 and 408, to
prevent net fluid movement along the separation channel during
injection. Power supplies capable of supplying the voltages and
currents required to implement this and other embodiments of the
invention are described in U.S. Pat. No. 5,965,001, which is
assigned to the assignee of the present invention.
[0044] As the sample is injected from channel 472 toward the
opposing wall of the "T" intersection between channels 472 and 404,
the sample accumulates in the portion of channel 404 near the
intersection, providing a more concentrated sample. The longer the
sample is concentrated in this manner, the more sample will be
available for separation and detection. The degree of concentration
appropriate for a given analysis will represent a simple trade-off
between loading time and sensitivity. A "high sensitivity" script
can be run on the same chip as a "normal" script, the only
difference being a longer loading time and therefore total analysis
time. By way of further example and not limitation, the techniques
described herein can be used in connection with the inventions
disclosed in Caliper U.S. Pat. Nos. 5,976,336 and 6,153,073, both
of which are assigned to the assignee of the present invention.
[0045] The increased conductivity of region 490 allows an
electrical potential to be maintained across the blind "T"
intersection, enabling the flow of fluid from channel 472 to be
directed into a wall. The increase in conductivity can be set based
on the separation between the channels between which the glass
resistor provides a conductive path, and the level of ion
implantation in the glass resistor. The amount ion implantation can
be determined by the dose amount, the energy level used during
implantation, and the thermal annealing process. In an exemplary
embodiment, the sheet conductivity of a glass substrate, as
measured by a four point probe, is about 10.sup.15 ohms per square
(.OMEGA./). Ion implantation can increase the glass conductivity by
several orders of magnitude, for example up to about 10.sup.8
.OMEGA./. Prawer et al., supra, shows that a sheet resistance of
10.sup.8 .OMEGA./ can be achieved by implanting carbon at a dose of
10.sup.16 ions/cm.sup.2. This is a high dose, but is attainable
using standard implantation methods known in the art. This level of
sheet resistance is suitable for the present invention, as can be
seen by the following example.
[0046] A microfluidic device similar to that shown in FIGS. 4 and 5
with 40 .mu.m wide channels and with a gap of 100 .mu.m between
channel 490 and channels 466 and 468 can be manufactured using the
methods described above. In this exemplary embodiment, the glass
resistor electrically connecting channel 490 and channel 466 and
468 could have a length to width ratio of about 2 (allowing for
some lateral current spreading). Assuming a sheet resistance of
10.sup.8 .OMEGA./, the resistance of the glass resistor would be
2.times.10.sup.8 ohms. If the current used to inject the sample
were about 3 .mu.A, which is typical of microfluidic devices in
accordance with the invention, the voltage drop across the glass
resistor would be 600V. This is an easily achievable value, since
the high voltage power supplies used in electrophoretic
microfluidic analysis systems typically supply voltages in the
range of 1500V to 3000V, as is the case with the Agilent 2100
Bioanalyzer, and the Caliper Technologies AMS-90 systems.
Exemplary Methods
[0047] Various methods can use microfluidic devices in accordance
with the present invention. Such methods include, but are not
limited to separating macromolecules by capillary electrophoresis
and detecting reactions. Such methods employ a microfluidic device
comprising an insulating substrate, at least one of a microchannel
and a well formed in the insulating substrate, and at least one ion
implanted region in the insulating substrate located at or adjacent
the at least one of the microchannel and the well, the at least one
ion implanted region having increased conductivity compared to the
insulating substrate.
[0048] A method of separating macromolecules by capillary
electrophoresis according an embodiment of the present invention
comprises: providing a microfluidic device, as described above;
introducing a sample containing the macromolecules into one end of
the microchannel; and applying a voltage gradient across the length
of the microchannel, whereby the macromolecules in the sample are
separated in the microchannel.
[0049] A method of detecting a reaction according an embodiment of
the present invention, comprises the steps of: introducing a first
reagent into a microchannel of the microfluidic device; introducing
a second reagent into the microchannel, whereby the first and
second reagents mix together to form a reagent mixture; introducing
a test compound into the reagent mixture; and detecting an effect
of the compound on the reagent mixture.
CONCLUSION
[0050] While various embodiments of the present invention have been
described above, it should be understood that they have been
presented by way of example, and not limitation. It will be
apparent to persons skilled in the relevant art that various
changes in form and detail can be made therein without departing
from the spirit and scope of the invention. This is especially true
in light of technology and terms within the relevant art(s) that
may be later developed. For example, the present invention is
applicable in a variety of applications, such as electrical
connectors for application of electrical energy, as in-line sensors
for sensing conductivity, temperature, or the like. Alternatively,
increased conductivity regions can be used on chips to apply or
modify electric fields for various purposes. By way of further
example and not limitation, the techniques described herein can be
used in connection with the inventions disclosed in Caliper U.S.
Pat. No. 5,965,410, which is assigned to the assignee of the
present invention.
[0051] The present invention has been described above with the aid
of functional building blocks, modules or steps illustrating the
performance of specified functions and relationships thereof. The
collection of sub-steps or boundaries of these functional building
blocks have been defined herein for the convenience of the
description. Alternate boundaries can be defined so long as the
specified functions and relationships thereof are appropriately
performed. Any such alternate collection of sub-steps or boundaries
are thus within the scope and spirit of the claimed invention.
Thus, the breadth and scope of the present invention should not be
limited by any of the above-described exemplary embodiments, but
should be defined only in accordance with the following claims and
their equivalents. 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.
* * * * *