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Description  |
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BACKGROUND OF THE INVENTION
Microfluidic devices and systems have been gaining substantial interest as
they are increasingly being demonstrated to be robust, highly accurate,
high throughput and low cost methods of performing previously cumbersome
and or expensive analytical operations.
In particular, microfluidic systems have been described for use in ultra
high throughput screening assay systems, e.g., for pharmaceutical
discovery, diagnostics and the like. See International Application No.
PCT/US97/10894 filed Jun. 28, 1997 (Attorney Docket No. 17646-000420PC).
In addition, such microfluidic systems have reportedly been used in
performing separations-based analyses, e.g., nucleic acid separations,
etc. See, e.g., Woolley et al., Proc. Nat'l Acad. Sci., USA 91:11348-11352
(1994).
Despite the promise of microfluidic systems in terms of throughput,
automatability and cost, many of the systems that have been described
suffer from substantial drawbacks. Initially, many of these systems have
substantial reductions in resolution over their counterpart methods on the
bench top. In particular, a number of relatively minor considerations can
readily become major factors when considered in the context of the
relatively small amounts of material transported through these systems.
For example, in microfluidic channels that include curves or turns,
variations in distances through these turns and curves at the inside and
outside edges can substantially affect the resolution of materials
transported through these channels.
Further, simple operations, such as dilution and mixing have generally been
accomplished at the expense of overall device volume, e.g., adding to the
reagent/material volume required for carrying out the overall function of
the device. In particular, such mixing typically requires much larger
chambers or channels in order to provide adequate mixing of reagents or
diluents within the confines f the microfluidic systems.
Thus, it would be generally desirable to provide microfluidic systems that
are capable of capitalizing upon the myriad benefits described above,
without sacrificing other attributes, such as resolution, volume, and the
like. The present invention meets these and other needs.
SUMMARY OF THE INVENTION
The present invention generally provides microfluidic devices, systems and
methods of using these devices and systems. The microfluidic devices and
systems generally incorporate improved channel profiles that result in
substantial benefits over previously described microfluidic systems.
For example, in one embodiment, the present invention provides microfluidic
devices and systems incorporating them, which devices comprise a body
structure and at least a first microscale channel disposed therein. The
microscale channel typically comprises at least first and second ends and
at least a portion of the microscale channel having an aspect ratio
(width/depth) less than 1. In preferred aspects, the devices and systems
include an electrical controller operably linked to the first and second
ends of the microscale channel, for applying a voltage gradient between
the first and second ends, and/or are fabricated from polymeric materials.
In a related but alternate embodiment, the present invention provides
microfluidic devices and systems that comprise a body structure having at
least a first microscale channel disposed therein, where the microscale
channel has at least one turning portion incorporated therein. In this
embodiment, the turning portion of the channel comprises a varied depth
across its width, where the varied depth is shallower at an outside edge
of the turning portion than at an inside edge of the turning portion.
Preferably, the relative depths at the inside edge and outside edge of the
turning portion of the channel are selected whereby the time required for
a material traveling through the turning portion at the outside edge is
substantially equivalent to a time required for the material to travel
through the turning portion at the inside edge.
As alluded to above, the present invention also comprises microfluidic
systems that include the above described microfluidic devices in
combination with an electrical control system. The electrical control
system is operably coupled to the first and second ends of the first and
second channels, and capable of concomitantly delivering a voltage to each
of the first and second ends of the first and second channels.
BRIEF DESCRIPTION OF THE FIGURES
FIG. 1 schematically illustrates a microfluidic device fabricated from a
planar substrate.
FIGS. 2A and 2B illustrate the distortion of material regions or plugs when
transported through a typical, curved microfluidic channel. FIG. 2A
illustrates distortion for a single material region, while FIG. 2B
illustrates distortion for multiple separate species regions or bands,
such as in electrophoretic separations analysis, as well as exemplary
signal, e.g., fluorescent signal that would be obtained from such species
bands, both before and after the distorting effects of the channel curves.
FIG. 3 illustrates a diagram of an electric field applied across the length
of a turning microscale channel.
FIG. 4 illustrates a comparison of a channel having a shallow aspect ratio,
e.g., >1 (width/depth)(top), as well as a channel having a deep aspect
ratio, e.g., <1 (width/depth)(bottom).
FIGS. 5A and 5B illustrate microscale channels having a varied depth to
permit improved material intermixing.
DETAILED DESCRIPTION OF THE INVENTION
I. General
A. Introduction
The present invention is generally directed to improved microfluidic
devices, systems and methods of using same, which incorporate channel
profiles that impart significant benefits over previously described
systems. In particular, the presently described devices and systems employ
channels having, at least in part, depths that are varied over those which
have been previously described. These varied channel depths provide
numerous beneficial and unexpected results.
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.
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. For example, typically, the body structure is fabricated from at
least two substrate layers that are mated together to define the channel
networks of the device, e.g., the interior portion. In preferred aspects,
the bottom portion of the device comprises a solid substrate that is
substantially planar in structure, and which has at least one
substantially flat upper surface.
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, reactive ion etching (RIE),
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.
In particularly preferred aspects, the substrate materials will 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 may include treated surfaces,
e.g., derivatized or coated surfaces, to enhance their utility in the
microfluidic system, e.g., provide enhanced fluid direction, e.g., as
described in U.S. patent application Ser. No. 08/843,212, filed Apr. 14,
1997, and which is incorporated herein by reference in its entirety for
all purposes.
An example of a microfluidic device fabricated from a planar substrate is
illustrated in FIG. 1. Briefly, the channels and/or chambers of the device
100 are typically fabricated into or upon a flat surface of a planar
substrate 102. The channels and/or chambers of the device may be
fabricated using a variety of methods whereby these channels and chambers
are defined between two opposing substrates. For example, the channels,
e.g., channels 104 and 106, and/or chambers of the microfluidic devices
are typically fabricated into the upper surface of the bottom substrate or
portion, as microscale grooves or indentations, using the above described
microfabrication techniques. Alternatively, raised regions may be
fabricated onto the planar surface of the bottom portion or substrate in
order to define the channels and/or chambers. The top portion or substrate
(not separately shown) also comprises a first planar surface, and a second
surface opposite the first planar surface. In the microfluidic devices
prepared in accordance with the methods described herein, the top portion
also includes a plurality of apertures, holes or ports 110-116 disposed
therethrough, e.g., from the first planar surface to the second surface
opposite the first planar surface.
The first planar surface of the top substrate is then mated, e.g., placed
into contact with, and bonded to the planar surface of the bottom
substrate, covering and sealing the grooves and/or indentations 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 110-116 in the top portion of the device are
oriented such that they are in communication with at least one of the
channels, e.g., 104 or 106, 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 110-116 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. As shown, channel
104, which serves as the main or analysis channel in the device shown,
intersects channel 106, which serves as a sample introduction channel, at
intersection 108. The analysis channel 104, also includes a serpentine
portion 118, which serves to extend the length of the analysis channel
without requiring substantially greater substrate area.
In many embodiments, the microfluidic devices will include an optical
detection window 120 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.
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, nucleic acid analysis, including 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, e.g., as
described in U.S. patent application Ser. No. 08/845,754, filed Apr. 25,
1997, and incorporated herein by reference. 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.
II. Channel Aspect Ratios
In a first aspect, the present invention provides microfluidic devices and
systems that comprise a body structure which has disposed therein, at
least one microscale channel or channel portion, which channel or channel
portion has an aspect ratio that is substantially the inverse of
previously described microscale channels.
In particular, microscale channels which have been described for use in
microfluidic systems, typically have employed channel dimensions in the
range of from about 50 to about 200 .mu.m wide and from about 5 to about
20 .mu.m deep. In any event, the aspect ratios of these channels
(width/depth) is well in excess of 2 and typically is in the range of from
about 7 to about 10. These aspect ratios have likely resulted, at least in
part, from the processes involved in the fabrication of microfluidic
systems, and particularly, the microscale channels incorporated therein.
Specifically, such channels are often fabricated in silicon based
substrates, such as glass, quartz, silicon, etc., using photolithographic
techniques. The chemistries involved in such techniques are readily used
to fabricate channels having widths and depths in the above-described
ranges. However, because these techniques involve etching processes, i.e.,
using generally available isotropic etching chemicals e.g., hydrofluoric
acid (HF), they are generally not as effective in producing channels
having depths substantially greater than those described, due to the
increased etching time required. Specifically, isotropic agents typically
etch uniformly in all directions on amorphous substrates, i.e., glass. In
such instances, it becomes effectively impossible to produce channels
having aspect ratios less than 1.
Microfluidic devices incorporating the above-described dimensions and
aspect ratios, have proven very useful in a wide variety of important
analytical applications. These applications include high-throughput
screening of pharmaceutical and other test compounds (See commonly
assigned U.S. application Ser. No. 08/761,575, filed Dec. 6, 1996),
nucleic acid analysis, manipulation and separation (See commonly assigned
U.S. application Ser. Nos. 08/835,101 and 08/845,754, filed Apr. 4, 1997
and Apr. 25, 1997, respectively), and more.
Despite the substantial utility of these systems, the present inventors
have identified some potential shortcomings of microscale channels having
the above-described dimensions, particularly where it is desired to
transport a given material region over a substantial distance within these
channels.
In a first particular example, because microfluidic systems are typically
fabricated within body structures that have relatively small areas, it is
generally desirable to maximize the use of the space within the body
structure. As such, channels often incorporate geometries that include a
number of channel turns or corners, e.g., serpentine, saw-tooth etc. The
incorporation of such channel turns can have adverse effects on the
ability of discrete material regions to maintain their cohesion. In
particular, in a turning microscale channel, material travelling along the
outside edge traverses the turn much more slowly than material travelling
at the inside edge, imitating a "race-track" effect. This effect is at
least in part due to the greater distance, or in the case of a three
dimension fluidic system, the greater volume a material must travel
through at the outside edge of a channel as opposed to the inner edge of
the channel. This difference in traversal time can result in a substantial
perturbation or distortion of a discrete, cohesive material plug or region
in a microscale channel.
A schematic illustration of this sample perturbation resulting from turns
or curves in microscale channels is provided in FIG. 2A. Briefly, FIG. 2A
illustrates a discrete material region 204, e.g., a sample plug, species
band or the like, travelling through a microscale channel 202. In the
straight portions of the channel, the material region substantially
maintains its shape with a certain level of diffusion. However, once the
material region travels around a curve in the channel, the differences in
flow rate through the channel at different points across the channel's
width, result in a distorted material region 206.
The distortion of these material regions can adversely effect the
resolution with which the particular material region is transported
through the turning channel. This is particularly problematic, for
example, in separation applications, e.g., protein or nucleic acid
electrophoresis, where the goal is to separate a given material into its
constituent elements, and to separately detect those elements. Further,
such separation applications typically require substantially longer
separation channels or columns, thereby increasing the number of channel
turns to which a particular material will be subjected. This separate
detection typically requires that the elements be maintained as well
resolved material regions. FIG. 2B illustrates an exemplary separation
channel incorporating a channel turn. The separated species bands 214
substantially maintain their shape and separation within the straight
portions of channel 202. The well-resolved character of the species bands
214 is illustrated by the signal graph 216. After having traveled through
the curved section of the channel, the species bands 218 show substantial
distortion. Further, as indicated by the signal graph 220, the resolution
of these bands, and thus their detectability, is substantially reduced.
This material region distortion, or sample perturbation, also becomes a
problem where one wishes to separately transport materials through a
microfluidic system, e.g., for separate analysis. Examples of such
applications include high-throughput experimentation, e.g., screening,
diagnostics, and the like. In particular, maintaining different materials,
e.g., samples, as well-resolved plugs of material, i.e., without the
above-described distortion effects, allows multiple plugs to be moved
through a system without fear of intermixing. This also prevents the
excess, uncontrolled dilution of those samples, resulting from the
distortion effects.
In certain preferred aspects, the microfluidic devices and systems
described herein, employ electrokinetic material transport systems for
moving and directing material through and among the microscale channel
networks that are incorporated in the microfluidic devices. Unfortunately,
however, the level of sample perturbation is substantially increased in
microfluidic systems that employ such electrokinetic material transport.
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, i.e., cations will move toward the
negative electrode, while anions will move toward the positive electrode.
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.
"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
allowing the right terminus to float (applying no voltage). 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 transpor | | |
