U.S. patent application number 11/628416 was filed with the patent office on 2008-05-15 for device for handling drops for biochemical analysis, method for producing said device and a system for microfluidic analysis.
Invention is credited to Francois Caron, Christian Druon, Jean-Christopher Fourrier, Pierre Tabourier.
Application Number | 20080110753 11/628416 |
Document ID | / |
Family ID | 34946857 |
Filed Date | 2008-05-15 |
United States Patent
Application |
20080110753 |
Kind Code |
A1 |
Fourrier; Jean-Christopher ;
et al. |
May 15, 2008 |
Device For Handling Drops For Biochemical Analysis, Method For
Producing Said Device And A System For Microfluidic Analysis
Abstract
A device for handling drops on a displacement by electrowetting
plane, including at least one displacement track. The track
includes an electrically insulating substrate on the surface of
which rest two or more interdigitated conducting electrodes. These
electrodes are covered by a dielectric insulating layer, itself
covered by a partially-wetting layer. Also, a method for the
manufacture of the aforementioned device, in which the creation of
the partially-wetting layer includes the creation of a mask in a
photosensitive material by the deposition of this material onto a
substrate, then a photolithographic stage, followed by development
of the photosensitive material, the deposition of a non-wetting
material onto the mask, at least one annealing process before
dissolution, dissolution of the mask, and at least one annealing
process after dissolution. Also, a system for the microfluidic
analysis of a liquid sample.
Inventors: |
Fourrier; Jean-Christopher;
(Montceau-Les-Mines, FR) ; Caron; Francois;
(Douai, FR) ; Tabourier; Pierre; (Calais, FR)
; Druon; Christian; (Villeneuve D'Ascq, FR) |
Correspondence
Address: |
LADAS & PARRY LLP
224 SOUTH MICHIGAN AVENUE, SUITE 1600
CHICAGO
IL
60604
US
|
Family ID: |
34946857 |
Appl. No.: |
11/628416 |
Filed: |
June 6, 2005 |
PCT Filed: |
June 6, 2005 |
PCT NO: |
PCT/FR05/01385 |
371 Date: |
August 20, 2007 |
Current U.S.
Class: |
204/403.01 ;
204/400; 427/2.11; 430/5 |
Current CPC
Class: |
F04B 19/006 20130101;
B01L 3/502707 20130101; B01L 2400/0427 20130101; B01L 3/502792
20130101; F05C 2225/04 20130101; B01L 2400/088 20130101; B01L
3/502746 20130101; B01L 2300/0887 20130101; B01L 2300/165
20130101 |
Class at
Publication: |
204/403.01 ;
427/2.11; 430/5; 204/400 |
International
Class: |
C25B 9/06 20060101
C25B009/06 |
Foreign Application Data
Date |
Code |
Application Number |
Jun 4, 2004 |
FR |
0406080 |
Claims
1. A device for handling drops on a displacement by electrowetting
plane, including at least one track, wherein said track comprises:
an electrically insulating substrate with a top surface, at least
two first conducting electrodes with a top surface and a bottom
surface, resting by their bottom surface on said top surface of the
said electrically insulating substrate, with each of the said first
electrodes being interdigitated with at least one other of these
said first electrodes, a dielectric insulating layer with a bottom
surface and a top surface, resting by its bottom surface on said
top surface of said first electrodes, a partially-wetting layer
with a bottom surface and a top surface, resting by its bottom
surface on the top surface of the said dielectric insulating
layer.
2. A device according to claim 1, further comprising at least one
counter-electrode separate from said first electrodes.
3. A device according to claim 2, wherein said separate
counter-electrode is an earth line located on or under the top
surface of, or inserted into, the said partially-wetting layer.
4. A device according to claim 1, further comprising a second track
positioned opposite to and separated from the first track, so that
a space is formed between said first and second tracks, with said
second track including a non-wetting layer with a bottom surface on
one side of said space and a top surface on the other side.
5. A device according to claim 4, wherein said non-wetting layer of
said second track is partially wetting.
6. A device according to claim 4, wherein said second track
includes a top layer that is electrically insulating,
semiconducting or conducting, located on one side of the top
surface of the said non-wetting layer.
7. A device according to claim 4, wherein said second track
includes one or more counter-electrodes located between said
non-wetting layer and the said top layer.
8. A device according to claim 7, wherein said second track
includes a dielectric insulating layer located between said
non-wetting layer and the said counter-electrode(s).
9. A device according to said partially-wetting layer of said first
track and/or of the said second track includes non-wetting zones
and wetting zones, with said wetting zones being reactive
functionalised zones.
10. A device for handling drops between two displacement by
electrowetting planes, including two tracks separated by a space,
the device comprising: the first track includes: an electrically
insulating substrate with a top surface, at least two first
electrodes with a top surface and a bottom surface, resting by
their bottom surface on said top surface of said electrically
insulating substrate, with each of said first electrodes being
interdigitated with at least one other of these said first
electrodes, a non-wetting layer with a bottom surface and a top
surface, located on one side of the top surface of the said first
electrodes, the second track includes: a partially-wetting layer
with a top surface and a bottom surface, where said
partially-wetting layer of the said first track and/or of said
second track includes non-wetting zones and wetting zones, and
where the said wetting zones are reactive functionalised zones.
11. A device according to claim 10, wherein said first track
includes a dielectric insulating layer located between the top
surface of the said first electrodes and the bottom surface of the
said non-wetting layer.
12. A device according to claim 10, further comprising an earth
line located on or under the top surface of, or inserted into, the
said non-wetting layer.
13. A device according to claim 10, wherein said second track
includes a layer that is electrically insulating, conducting or
semiconducting, located on one side of the top surface of the said
non-wetting layer.
14. A device according to claim 1, wherein said electrically
insulating substrate of said first track is transparent.
15. A device according to claim 14, wherein said electrically
insulating substrate of said first track is a glass substrate.
16. A device according to claim 9, wherein said wetting zones are
openings in non-wetting zones.
17. A device according to claim 9, wherein said wetting zones are
biochemically functionalised and reactive.
18. A device according to claim 1, wherein said non-wetting layer
and/or said non-wetting zones of said partially-wetting layer, are
non-wetting in relation to water and therefore hydrophobic, and
said wetting zones are wetting in relation to water and therefore
hydrophilic.
19. A device according to claim 1, wherein said non-wetting layer
and/or said non-wetting zones of the said partially-wetting layer
are in tetrafluoroethylene polymer.
20. A method for the manufacture of the device according to claim
1, in which the creation of said partially-wetting layer of said
first track or the said second track includes: a stage for the
creation of a mask in a photosensitive material, by deposition of
the said photosensitive material onto a substrate, then
photolithography, and then development of the said photosensitive
material, a stage for the deposition of a non-wetting material onto
the said mask, at least one annealing stage before dissolution, a
stage for the dissolution of the said mask, at least one annealing
stage after dissolution.
21. A method according to claim 20, wherein the annealing
temperature of said annealing stage before dissolution is lower
than the annealing temperature of said annealing stage after
dissolution.
22. A method according to claim 20, wherein said stage for the
deposition of a non-wetting material onto said mask is a stage for
the deposition of a tetrafluoroethyene polymer.
23. A system for the microfluidic analysis of a liquid sample,
characterised in that it includes: at least one means for preparing
the liquid sample with at least one outlet, at least one drop
handling device according to claim 1, coupled by one of its inlets
to one of the outlets of the said preparation means, and with at
least one outlet, at least one analysis means coupled by one of its
inlets to one of the outlets of the said drop handling device.
24. A system according to claim 23, wherein said preparation means
includes one or more loading reservoirs or docks.
25. A system according to claim 23, wherein said analysis means is
a mass spectrometer, a fluorescence detector, or a UV light
detector.
26. A system according to claim 23, wherein the system is
integrated into a microlaboratory.
27. A device according to claim 15, wherein said wetting zones are
openings in non-wetting zones.
28. A device according to claim 27, wherein said wetting zones are
biochemically functionalised and reactive.
29. A device according to claim 28, wherein said non-wetting layer
and/or said non-wetting zones of said partially-wetting layer, are
non-wetting in relation to water and therefore hydrophobic, and
said wetting zones are wetting in relation to water and therefore
hydrophilic.
30. A device according to claim 29, wherein said non-wetting layer
and/or said non-wetting zones of the said partially-wetting layer
are in tetrafluoroethylene polymer.
31. A method for the manufacture of the device according to claim
10, in which the creation of said partially-wetting layer of said
first track or the said second track includes: a stage for the
creation of a mask in a photosensitive material, by deposition of
the said photosensitive material onto a substrate, then
photolithography, and then development of the said photosensitive
material, a stage for the deposition of a non-setting material onto
the said mask, at least one annealing stage before dissolution, a
stage for the dissolution of the said mask, at least one annealing
stage after dissolution.
32. A method according to claim 31, wherein the annealing
temperature of said annealing stage before dissolution is lower
than the annealing temperature of said annealing stage after
dissolution.
33. A method according to claim 31, wherein said stage for the
deposition of a non-wetting material onto said mask is a stage for
the deposition of a tetrafluoroethyene polymer.
34. A system for the microfluidic analysis of a liquid sample,
characterised in that it includes: at least one means for preparing
the liquid sample with at least one outlet, at least one drop
handling device according to claim 10, coupled by one of its inlets
to one of the outlets of the said preparation means, and with at
least one outlet, at least one analysis means coupled by one of its
inlets to one of the outlets of the said drop handling device.
35. A system according to claim 34, wherein said preparation means
includes one or more loading reservoirs or docks.
36. A system according to claim 34, wherein said analysis means is
a mass spectrometer, a fluorescence detector, or a UV light
detector.
37. A system according to claim 34, wherein the system is
integrated into a microlaboratory.
Description
[0001] The subject of this present invention is a drop handling
device that is intended for biochemical analysis, a method for the
manufacture of such a device, and a microfluidic analysis system
using such a device.
[0002] Nowadays, the new technologies allow the design of systems
of micrometric and nanometric dimensions and with very high levels
of complexity. Ideally, these systems come with all sorts of
features, and are used in many areas such as biology or
biochemistry. In particular, protoemics, an activity associated
with the identification and the study of proteins, attempts to use
the new technologies to reduce the sampled volumes being
manipulated, and to reduce contamination. Generally speaking, the
objective is to control the microhandling of the material, before
spectrometric analysis for example.
[0003] In such microsystems, the problem of controlling the fluid
flows arises from a strategic viewpoint, to the extent that the
material, such as proteins for example, cannot be manipulated other
than in a liquid medium. The invention therefore relates to the
area of microfluidics, which more generally concerns flows in
systems of micrometric or nanometric dimensions, in which the
manipulated sample can subjected to electric fields or to
partitioning effects of a complex physical or chemical nature, and
in which the high area/volume ratio is very important.
[0004] In this area, the reduction in the size of the systems
results in a reduction in the volumes and the reaction times or in
shorter exchanges, and the ability to integrate several modules
with different features such as transportation, treatment, or
indeed analysis, all on a single wafer of silicon for example.
[0005] In order to transport the liquid, two types of fluidic
displacement are generally possible, namely the pumping of a
continuous flow, and the displacement of calibrated microvolumes.
The displacement of calibrated microvolumes has a certain number of
advantages. In fact, it allows very small liquid volumes and allows
an appropriate control of the flow of the microvolumes, while
continuous-flow pumping is characterised by a constant flow. In
addition, this type of displacement allows a variety of
synchronisations that allow mixing of the liquids for example. In
order to achieve fluidic displacement of the displacement of
calibrated microvolumes type, different methods of operation are
known, such as by pneumatic action, by acoustic surface waves, by
dielectrophoretic effect, by electrowetting, and by electrowetting
on dielectric (EWOD). This last a method makes use of a relatively
simple technological system and allows control of the flow and the
circulation of a calibrated volume of conducting liquid in a
network of electrodes.
[0006] American patent U.S. Pat. No. 6,565,727, and the publication
by Cho et al of "Particle separation and concentration control for
digital microfluidic systems", are known in particular, describing
the displacement of drops by electrowetting, as described above.
However, the devices described in these publications have a bottom
part that includes electrodes and a top part that includes
counter-electrodes, with the drop moving between these parts. This
top part in particular renders the device more bulky and more
complex.
[0007] In addition, the manipulated samples are often very precious
and in very small quantities. There is therefore a requirement to
optimise handling of these samples, by chemical treatment or by
interacting with the material during transportation. The known
microfluidic displacement systems, whether they necessitate two
facing substrates or a single substrate, and whether they use a
counter-electrode or not, do not permit this optimisation. In fact,
in particular in the publication of Cho et al. entitled "Particle
separation and concentration control for digital microfluidic
systems", a device is proposed which allows interaction with the
drop physically, by direct interaction between the electrodes and
the drop during transportation, and not chemical interaction. This
chemical interaction, which is necessary for optimising handling of
the samples, is therefore impossible in the device of Cho et
al.
[0008] In fact, this optimisation is rendered extremely difficult
by the fact that the displacement requires one or more tracks in
hydrophobic materials in order to limit friction and hysteresis in
the displacements. This hydrophobic character of the displacement
track in particular prevents chemical treatment of, or interaction
with, the material during transportation.
[0009] It should be noted here that one is generally more
interested in the non-wettability property of the displacement
track in relation to any given liquid. When the liquid is aqueous,
as is generally the case when one is handling proteins for example,
the non-wettability and the wettability in relation to water are
the properties of hydrophobicity and hydrophilicity respectively. A
hydrophobic material is a non-wetting material in relation to
water, and a hydrophilic material is a wetting material in relation
to water. Wettability is generally characterised by the angle
(.theta.) of contact between the drop (1) and the surface (2) (see
FIGS. 1a to 1d). Use is sometimes made of the wettability
coefficient, defined as the cosine of the aforementioned angle.
Perfect wettability thus corresponds to a wettability coefficient
of 1, and so to .theta.=0.degree.. Total absence of wettability
then corresponds to a wettability coefficient of -1 (minus 1), and
so to .theta.=180.degree.. In what follows, we will therefore speak
of wetting material in relation to a liquid for a material whose
wettability coefficient in relation to this liquid tends to 1
(without necessarily being equal to 1), as illustrated in FIG. 1a,
and we will speak of non-wetting material in relation to a liquid
for a material whose wettability coefficient in relation to this
liquid tends to -1 (without necessarily being equal to -1) as
illustrated in FIG. 1b. FIGS. 1c and 1d illustrate intermediate
cases of wettability (.theta.<90.degree.) or non-wettability
(.theta.>90.degree.) respectively.
[0010] The problem posed by the non-wetting materials in relation
to a liquid, in particular the hydrophobic materials, also
essential to the displacement, is that the surface properties of
these materials prevent the creation of surface chemical treatment
zones due to the fact that these materials are characterised by a
low surface energy. If one tries to functionalise the surface of
such materials locally, which would allow chemical treatment of the
manipulated liquids, the result is not very reliable, difficult to
control and too imperfect. The alternative, consisting of rendering
the non-wetting material more rigorous in relation to the liquid,
is not an option since it eliminates the ability of the material to
favour the transportation of the liquid. It is therefore necessary
to use a layer of material which is partially wetting, meaning that
it is necessary to maintain the non-wetting character for the
displacement, while also creating wetting zones or zones of high
wettability for the functionalisation.
[0011] Applied to the particular case where the material concerned
is hydrophobic, one is particularly aware of two conventional
photolithographic techniques for the creation of a partially
hydrophobic layer by the creation of openings in a hydrophobic
material, where these openings become hydrophilic zones distributed
in the hydrophobic layer. In a first technique (FIG. 5), also used
by Cho et al. in "Particle separation and concentration control for
digital microfluidic systems", after the deposition of a layer of
hydrophobic material onto a substrate, a layer of photosensitive
resin is deposited which contains a surface-active agent, a
chemical substance used to increase the wettability of a surface in
relation to a liquid. This technique in particular poses the
problem of definitive pollution of the hydrophobic material and
therefore loss of the ability of this material to favour the
displacement of a liquid. In the second technique (FIG. 6), after
deposition of a layer of hydrophobic material onto a substrate, and
before deposition of a photosensitive resin, the layer of
hydrophobic material is first subjected to surface modification by
means of a plasma, in order to alter its hydrophobic properties,
meaning to render it less hydrophobic. This technique also poses
the problem of definitive alteration of the surface properties of
the hydrophobic material.
[0012] With such techniques, either the openings created, and
therefore the hydrophilic zones, are not sufficiently distinct and
precise, possible with hydrophobic deposits, and as a consequence
unsuitable for the creation of chemically functionalised zones, or
the hydrophobic zones will have their properties modified and their
hydrophobic character diminished, with consequent unsuitability for
the displacement of liquid. The same comments apply in the case of
application of these techniques for the creation of wetting zones
in a layer that is non-wetting in relation to the liquid
transported.
[0013] There is therefore a requirement for a method which can be
used to render a non-wetting transportation track partially wetting
in relation to the liquid transported, and in particular partially
hydrophilic when the liquid is a solution containing water, so that
the ability to transport the drop of liquid is maintained, while
also allowing chemical treatment or interaction with this drop
during transportation.
[0014] More generally, there exists the need for a reliable
solution which can be used to overcome the aforementioned
drawbacks, in particular the optimisation of displacement and the
manufacture of an optimised displacement track.
[0015] The purpose of the invention is therefore to overcome these
drawbacks. To this end, the invention relates, according to a first
aspect, to a device for handling of a drop in a displacement by
electrowetting plane, which includes at least one displacement by
electrowetting track, and which allows chemical treatment of, or
interaction with, the drop simultaneously with its
transportation.
[0016] The displacement track includes at least two interdigitated
electrodes which rest on an electrically insulating substrate and
which are covered by an insulating dielectric layer. This assembly
of insulating substrate, electrodes, and insulating dielectric
layer, is covered with a layer that is partially wetting in
relation to the manipulated drops.
[0017] In an implementation variant concerning handling of drops
containing water, the partially-wetting layer is therefore a
partially-hydrophilic layer.
[0018] In the remainder of the description, and in order to
simplify the description, we will speak of layers or materials that
are respectively non-wetting, partially-wetting, or wetting, to
mean layers or materials that are respectively non-wetting,
partially-wetting, or wetting in relation to the manipulated
drops.
[0019] In another implementation variant, the device of the
invention includes at least one counter-electrode which is separate
from the first electrodes. This counter-electrode can be an earth
line which will then be located on, under or in the
partially-wetting layer.
[0020] In an implementation variant, possibly in combination with
the preceding one, the device includes a second track positioned
opposite to and separated from the first track, so that a space,
intended to be filled by an electrically insulating fluid that is
non-miscible in relation to the drop transported, is formed between
the first and second tracks, with the second track including a
non-wetting layer directly in contact with the space thus formed.
This non-wetting layer of the second track can possibly be
partially wetting. This non-wetting layer is also possibly covered
by a top layer which is either electrically insulating,
semiconducting, or conducting.
[0021] In another implementation variant, the second track includes
one or more counter-electrodes located between the non-wetting
layer and the top layer. It can also possibly include an insulating
dielectric layer which will be located between the said non-wetting
layer and the said counter-electrode(s).
[0022] Possibly in combination with each of these implementation
variants of the device, the partially-wetting layer of the first
track and/or of the second track includes non-wetting zones and
wetting zones, where the wetting zones are reactive functionalised
zones.
[0023] In another implementation variant, the device of the
invention for handling a drop in a plane includes two tracks
separated by a space that is intended to be filled by an
electrically insulating fluid which is non-miscible in relation to
the drop transported. The first track includes a layer or
electrically insulating substrate on which rests at least two
interdigitated electrodes. On this assembly rests a non-wetting
layer. The second track includes a partially-wetting layer. The
partially-wetting layer of the first track and/or of the second
track includes non-wetting zones and wetting zones, where the
wetting zones are reactive functionalised zones.
[0024] In this implementation variant, the first track can possibly
also include an insulating dielectric layer located between the
electrodes and the non-wetting layer. Possibly also, the device in
this implementation variant includes an earth line located on,
under or inserted into the non-wetting layer.
[0025] In an implementation variant, the second track includes a
top layer which is electrically insulating, semiconducting, or
conducting.
[0026] In combination with each of these implementation variants of
the device, the electrically insulating substrate of the first
track is preferably transparent, like a glass substrate for
example.
[0027] In one or more of the preceding variants, the wetting zones
are preferably biochemically functionalised and reactive.
[0028] These wetting zones are preferably openings in non-wetting
zones. The non-wetting material constituting the non-wetting layer
and/or the non-wetting zones of the partially-wetting layer, is
preferably a tetrafluoroethyene polymer.
[0029] Thus, the device of the invention advantageously allows
handling of a drop of liquid, by transporting it in a plane by
electrowetting, on a single track or between two facing tracks,
with or without the use of a counter-electrode, while also acting
chemically on the drop during its passage through chemically
functionalised zones. The desired optimisation is therefore
achieved, namely reducing the preparatory treatments to a later
analysis in a microsystem, during transportation, in order to avoid
contamination and the loss of samples that are very costly and in
very small volumes, while also allowing for the aforementioned
constraints of microfluidics.
[0030] According to a second aspect, the invention relates to a
method for the manufacture of the aforementioned device, in which
creation of the partially-wetting layer of the first or of the
second track is derived from the technique known as "lift off",
used in microelectronics to create patterns in metal. Although this
"lift off" technique, as it is commonly known, allows deposition of
the non-wetting layer at the last stage, thus avoiding a
prejudicial surface treatment, it is not suitable however for the
creation of patterns in such a non-wetting material, in particular
a hydrophobic material such as a tetrafluoroethyene polymer, since
it does not allow the creation of wetting zones that are distinct
and precise in this non-wetting material. The invention therefore
relates, according to this second aspect, to a method for the
manufacture of the aforementioned device, in which the creation of
the partially-wetting layer of the first or of the second track
includes the following stages: creation of a mask in photosensitive
material by deposition of the photosensitive material onto a
substrate, followed by photolithography, and then development of
the photosensitive material, deposition of a non-wetting material
onto the mask, at least one annealing process before dissolution,
dissolution of the mask, and at least one annealing process after
dissolution.
[0031] In an implementation variant, the temperature of the
annealing process before dissolution is lower than the temperature
of the annealing process after dissolution.
[0032] In another implementation variant, the first annealing
process before dissolution is followed by at least one other
annealing process at a temperature above that of the first
annealing process.
[0033] In another variant, possibly in combination with the
preceding one, the first annealing process after dissolution is
followed by at least one other annealing process at a temperature
above that of the first annealing process.
[0034] The dissolution of the mask can possibly be followed by
rinsing.
[0035] In another implementation variant, the non-wetting material
deposited is a tetrafluoroethyene polymer.
[0036] Thus, the method of the invention advantageously allows the
creation of a partially-wetting layer which contains wetting zones
that are distinct and precise, suitable for chemical
functionalisation, and which contains non-wetting zones that retain
their enhanced properties of non-wettability, which is necessary
for the transportation of drops. In fact, the layer of non-wetting
material is deposited at the last stage, is subjected to no surface
treatment, and is therefore subjected to no alteration of its
surface properties.
[0037] The invention relates finally, according to a third aspect,
to a system for the microfluidic analysis of a sample liquid, which
includes at least one means for preparing the sample, coupled to at
least one drop handling device according to the invention and, as
mentioned, itself coupled to at least one analysis means .
[0038] The preparation means preferably includes one or more
loading reservoirs or docks.
[0039] The analysis means is also preferably a mass spectrometer, a
fluorescence detector, or a detector of UV or IR emissions.
[0040] The system according to the invention can possibly be
integrated into a microsystem which itself includes one or more
laboratory operations that are usually performed manually, and
which will be known as microlaboratory operations.
[0041] Thus the system according to the invention advantageously
allows analysis of the liquid samples, after first preparing the
samples and then transporting them by the displacement of
calibrated microvolumes to an analyser, by automation of the
preparation and transportation tasks, built into a microlaboratory.
It therefore advantageously allows the risks of contamination and
loss of the material of the sample to be reduced, as well as
reducing the reaction times.
[0042] Other characteristics and advantages of the invention will
appear more clearly and more completely on reading the description
that follows of the preferred implementation variants of the method
and for creation of the device, provided by way of non-limiting
examples and with reference to the following appended drawings:
[0043] FIGS. 1a to 1d schematically illustrate the non-wettability
or wettability property of a surface in relation to a drop,
[0044] FIGS. 2a to 2r schematically represent different
implementation variants of the device according to the invention
(seen in section perpendicular to the direction of displacement of
the drop),
[0045] FIG. 3 schematically represents the displacement of a drop
on a track of the device according to a first implementation
variant,
[0046] FIG. 4 schematically represents the displacement of a drop
on a track of the device according to a second implementation
variant,
[0047] FIG. 5 schematically represents the method for the creation
of openings in a non-wetting material according to the conventional
photolithographic technique using a surface-active agent in
resin,
[0048] FIG. 6 schematically represents the method for the creation
of openings in a non-wetting material according to the conventional
photolithographic technique with surface alteration by plasma,
[0049] FIG. 7 schematically represents an implementation variant of
the method for the creation of openings in a non-wetting material
according to the invention,
[0050] FIG. 8 schematically illustrates the chemical
functionalisation of a wetting zone,
[0051] FIG. 9 schematically illustrates the chemical treatment of a
drop of a sample during its displacement,
[0052] FIG. 10 schematically represents an implementation variant
of the system according to the invention.
[0053] FIGS. 2a to 2r schematically represent different
implementation variants of the device of the invention (seen in
section perpendicular to the direction of displacement of the
drop).
[0054] In these FIGS. 2a to 2n, the device includes at least one
track with a substrate 1, preferably but not necessarily
transparent, in Pyrex.RTM. for example. Above this substrate 1 are
located the interdigitated electrodes 2. The notion of
interdigitated electrodes will be described later in greater detail
with reference to FIGS. 3 and 4.
[0055] On these electrodes 2 lies an insulating dielectric layer 3,
composed of oxides or polymers for example On this electrically
isolating layer 3 lies a non-wetting layer 4, which is rendered
partially wetting by the method for the creation of wetting
openings 5 in the non-wetting material 4. This method will be
described in more detail a little later with reference to FIG.
7.
[0056] In the implementation variants of FIGS. 2a to 2d, the device
includes a single track composed of layers 1, 2, 3 and 4. The
device of FIG. 2a allows the execution of a displacement by
electrowetting that does not require counter-electrodes. This
displacement will be explained later with reference to FIG. 3. The
devices of FIG. 2b each have a counter-electrode in the form of an
earth line 6 placed on the partially-wetting layer 4 (in FIG. 2b),
inserted into and not covered by the partially-wetting layer 4 (in
FIG. 2c), or inserted into and covered by the partially-wetting
layer 4 (in FIG. 2d) . For their part, the devices of FIGS. 2b to
2d allow execution of the displacement by electrowetting, with an
earth line as the counter-electrode. This displacement will be
described later with reference to FIG. 4.
[0057] FIG. 2e and those that follow show implementation variants
in which a second track is added, formed from a non-wetting layer 7
which itself is covered by a top layer 8 that can be either
electrically insulating or electrically semiconducting or indeed
electrically conducting. This second track is positioned in
relation to the first, with the use of spacers 9 employed to
maintain a displacement space 10 that is intended to be filled with
an electrically insulating fluid that is non-miscible in relation
to the drop transported.
[0058] It will be noted that, in order to achieve displacement by
electrowetting, the fluid filling the space 10 must actually be
electrically insulating. In addition, in order not to interact with
the drop transported, the fluid must actually be non-miscible in
relation to the liquid. It can be air or oil for example, in the
case of a drop of aqueous solution.
[0059] In particular, FIGS. 2f to 2h show implementation variants
which are respectively based on the devices of FIGS. 2b to 2d, to
which a second track is added as described previously.
[0060] In the implementation variant of the device of FIG. 2i, the
second track also includes one or more counter-electrodes 11
inserted between the non-wetting layer 7 and the top layer 8. There
is therefore no earth line, in contrast to the devices of FIGS. 2f
to 2h, since the counter-electrode is present in the second track.
The displacement method is nevertheless identical to that of FIGS.
2f to 2h.
[0061] The implementation variants of FIGS. 2j to 2l (seen in
section perpendicular to the direction of displacement of the drop)
are derived respectively and directly from the implementation
variants of FIGS. 2f to 2h, the only difference being that the
non-wetting layer 7 of the second track is rendered partially
wetting by the method for the creation of wetting openings 5 in the
non-wetting material 7, and which will be described later with
reference to FIG. 7.
[0062] FIG. 2m describes an implementation variant which is based
on that previously described in FIG. 2e, the only difference being
that the non-wetting layer 7 of the second track is rendered
partially wetting by the method for the creation of wetting
openings 5 in the non-wetting material 7, and which will be
described later with reference to FIG. 7.
[0063] For its part, the implementation variant of FIG. 2n is
derived from the implementation variant of FIG. 2i, the only two
differences being that the non-wetting layer 7 of the second track
is rendered partially wetting by the creation of wetting openings 5
in the non-wetting layer 7 according to the method which will be
described later with reference to FIG. 7, and in order to allow the
biochemical functionalisation of these wetting openings 5 without
interactions with the counter-electrode(s) 11, an insulating
dielectric layer 12 similar to that present in the first track is
inserted between the partially-wetting layer 7 and the
counter-electrode(s) 11.
[0064] The implementation variant described in FIG. 2o concerns a
device with two tracks. The first track differs from the first
track of the previous implementation variants in that the
non-wetting layer 4 of which it is composed is not partially
wetting, so that no wetting opening is created in this non-wetting
layer 4. In addition, this implementation variant requires no
insulating dielectric layer between the interdigitated electrodes 2
and the non-wetting layer 4 in the case where this non-wetting
layer 4 is itself electrically insulating. This is the case in
particular for a hydrophobic layer, in a material such as a
tetrafluoroethylene polymer. In practice however, such a material
is actually electrically insulating only if the thickness of the
layer is substantial (a thickness of the order of a micrometre). In
addition, in the case of FIG. 2o where the thickness of the
non-wetting layer 4 is not sufficient, it is possible to insert,
between the layer of interdigitated electrodes 2 and the
non-wetting layer 4, an insulating dielectric layer of the same
type as the layer 3 in the other figures.
[0065] On the non-wetting layer 4, there is located an earth line 6
acting as a counter-electrode. In this implementation variant, the
second track is identical to that of the implementation variants of
FIGS. 2j to 2m.
[0066] In the respective implementation variants of FIGS. 2p and
2q, the non-wetting layer 4 is not partially wetting since it has
no openings (5). These implementation variants are therefore
derived respectively from the variants of FIGS. 2k and 2l, with the
aforementioned difference (layer 4 totally non-wetting, while in
the variants of FIGS. 2k and 2l, it is partially wetting).
[0067] Finally, the implementation variant of FIG. 2r again uses
the displacement method of FIGS. 2a, 2e and 2m, meaning without the
use of a counter-electrode, and, as in the variants of FIGS. 2o to
2p, has a non-wetting layer 7 in the second track which is
partially wetting with the presence of the wetting openings 5, and
a non-wetting layer 4 in the first track which is totally
non-wetting since it has no wetting opening. In addition, as in the
variant of FIG. 2o, this implementation variant requires no
insulating dielectric layer between the interdigitated electrodes 2
and the non-wetting layer 4 in the case where this non-wetting
layer 4 is itself electrically insulating, this being the case in
particular for a hydrophobic layer, in a material such as a
tetrafluoroethylene polymer. Here again however, in practice, such
a material is actually electrically insulating only if the
thickness of the layer is substantial (a thickness of the order of
a micrometre) . In addition, in the case of FIG. 2r where the
thickness of the non-wetting layer 4 is not sufficient, it is
possible to insert, between the layer of interdigitated electrodes
2 and the non-wetting layer 4, an insulating dielectric layer of
the same type as layer 3 in the other figures.
[0068] FIG. 3 schematically represents the displacement of a drop
on a track of the device according to an implementation variant.
This figure breaks down into two parts. In the top part (diagrams
A, B and C), in the interests of simplification and so as to
facilitate the explanation, the representation of the device is a
representation from above and partial in that it shows neither the
non-wetting or partially wetting layer nor the dielectric
insulating layer, located between the drop (15) and electrodes 1,
2, 3 and 4. In the bottom part (diagrams A', B' and C'), the
representation of the device is a representation in section from
the side, in the direction of displacement of the drop.
[0069] More precisely, the device is of the same type as that of
FIG. 2a, meaning with a single track. However, the following
explanations concerning the displacement of the drop are applicable
more generally to the cases of FIGS. 2a, 2e, 2m and 2r, meaning
displacement on a track with interdigitated electrodes, without
counter-electrodes, and possibly with a second upper plane.
[0070] The device therefore requires several interdigitated
electrodes (1, 2, 3, 4) which rest on an electrically insulating
substrate 10 that is possibly transparent. On the layer of
interdigitated electrodes is located a dielectric insulating layer
11 and a non-wetting layer 12. This non-wetting layer 12 can be
partially wetting according to the configuration in which one finds
oneself (see FIG. 2 concerned), which does not alter the following
explanations concerning the displacement. The drop 15 is initially
on electrode 2 (stage A). By creating a potential difference
between electrode 3 and electrodes 1, 2 and 4, the drop moves onto
electrode 3 (stage B). In order to move it onto electrode 4, a
potential difference is created between electrode 4 and electrodes
1, 2 and 3. And so on.
[0071] FIG. 4 schematically represents the displacement of a drop
on a track of the device according to another implementation
variant. Here again, the figure breaks down into two parts. In the
top part (diagrams A, B and C), again in the interests of
simplification and in order to facilitate explanation as for FIG.
3, the representation of the device is a representation from above,
and partial in that it shows neither the non-wetting or partially
wetting layer nor the dielectric insulating layer, located between
the drop 15 and electrodes 1, 2, 3 and 4. In the bottom part
(diagrams A', B' and C'), the representation of the device is a
representation in section from the side, in the direction of
displacement of the drop.
[0072] More precisely, the device presented corresponds to a device
with a single track and an earth line as the counter-electrode, as
previously described at FIG. 2b. However, the following
explanations concerning the displacement of a drop on this device
are also applicable to the cases of FIGS. 2c, 2d, 2f, 2g, 2h, 2j,
2k, 2l, 2o, 2p, and 2q.
[0073] The device includes a layer of interdigitated electrodes (1,
2, 3, 4) which rest on an electrically and possibly transparent
insulating substrate 10. Above this layer of electrodes lies a
dielectric insulating layer 11. Above this dielectric insulating
layer 11 lies a non-wetting layer 12. This layer is possibly
partially wetting, depending on the configuration in which one
finds oneself (see FIG. 2). Above this non-wetting layer 12 (which
is possibly partially wetting) is located an earth electrode earth
line.
[0074] The drop 15 is initially on electrode 2 (stage A). By
creating a potential difference between electrode 3 and electrodes
1, 2, and 4 and the earth electrode, the drop moves onto electrode
3 (stage B). In order to move the drop onto the electrode 4, a
potential difference is created between electrode 4 and electrodes
1, 2, and 3 and the earth electrode, and so on.
[0075] If the earth electrode or earth line is replaced by a
counter-electrode located in an upper plane (the case of FIGS. 4i
and 4n), the previous explanations concerning FIG. 4 still
apply.
[0076] The method used to render partially wetting the non-wetting
layer of one of the tracks of the device of the invention, will now
be described with reference to FIG. 7, with a reminder of previous
designs with reference to FIGS. 5 and 6.
[0077] FIG. 5 schematically represents the stages of the method for
the creation of an opening in a non-wetting material, thus
rendering it partially-wetting, using the conventional
photolithographic technique with a surface-active agent. At stage
(a), a layer of non-wetting material 2 is deposited on a substrate
1. At stage (b), a layer of resin 3 containing a surface-active
agent is deposited on the non-wetting layer 2. The surface-active
agent is used to increase the wettability of the non-wetting layer
in relation to the resin, and therefore the adherence of the resin
to this layer. At stage (c), the photolithographic stage proper,
layer 3 is subjected to UV radiation. If layer 3 is in resin said
to be positive, then the ultraviolet radiation leads to rupturing
of the macromolecules of the exposed zones, conferring on these
zones increased solubility to the development solvent that will be
used at stage (d), while in contrast, the non-insulated parts will
be polymerised. This is therefore what happens, with the result of
the development stage (d). The development of the resin is
accompanied by an attack on the exposed non-wetting material and
therefore the appearance of zones or openings 4 in the non-wetting
layer 2 (stage (e)). This technique can be accompanied by
definitive alteration of the surface properties of the non-wetting
material due to the use of the surface-active agent in resin.
[0078] FIG. 6 schematically represents the stages of the method for
the creation of openings in a non-wetting material using the
conventional photolithographic plasma technique. This technique
differs from the preceding one in that it includes a complementary
stage that consists of subjecting the non-wetting layer 2 to
plasma-argon radiation (stage (b)) before deposition of the resin
layer. It is this radiation that will alter the surface properties
of the non-wetting layer 2, while in the preceding technique (FIG.
5), it is the presence of the surface-active agent in resin that
plays this role. The following stages (c), (d), (e), and (f)) are
respectively the same as stages (b), (c), (d) and (e) of FIG. 5.
The conclusion is the same as that for the conventional
photolithographic technique with a surface-active agent, namely
that there can be definitive alteration of the surface properties
of the non-wetting layer 2.
[0079] The method of the invention, now described with reference to
FIG. 7, is therefore a method for the manufacture of one or more
tracks of the device described previously, in which the creation of
the partially-wetting layer first includes a stage for the creation
of a mask in photosensitive material by the deposition of a layer
of this material 2 onto a substrate 1 (stage (a)), then
photolithography (stage (b)), and development of the photosensitive
material (stage (c)). In the implementation variant described in
FIG. 7, a negative resin is used as the photosensitive material,
meaning one in which the UV radiation leads to a polymerisation of
the insulated zones, leading to increased solubility of the zones
not exposed in the developer. It is therefore the zones not exposed
at stage (b) that disappear at stage (c), while the zones insulated
at stage (b) remain present at stage (c) and are identified by the
number 2. The choice of a negative resin does not limit the
invention in any way. The functioning of the method of the
invention is exactly the same with the use of a positive resin.
[0080] The stage (c) is followed by stage (d) for the deposition of
a layer of non-wetting material 3.
[0081] By way of an example, for the photolithographic stage, it is
possible to use a resin with the following parameters: [0082] resin
AZ 4562, [0083] development in AZ 351 B.
[0084] Stage (d) for deposition of the non-wetting material 3 is
followed by a first annealing stage. Depending the material chosen
(tetrafluoroethylene polymer for example), the annealing process
can be at 50.degree. C., and can last for 5 minutes. Preferably,
but not necessarily, this annealing process is followed by another
complementary annealing process. This second annealing process can
then be performed at a temperature of 110.degree. C., also for 5
minutes.
[0085] In the particular case of a hydrophobic material such as a
tetrafluoroethylene polymer, very little solvent remains in the
material at this stage. However a second annealing stage will be
needed after dissolution of the resin mask 2 (stage (e). In fact,
at the annealing temperatures of the hydrophobic material, the
resin polymerises, thus rendering it difficult to remove. The
consequence of this can be to leave traces of resin on the
substrate. These traces may well be difficult or even impossible to
remove during the following dissolution stage, and this can alter
the surface properties of the partially-wetting layer (partially
hydrophilic in the case of wettability in relation to water). The
openings may well not be perfectly non-wetting (or hydrophobic for
non-wettability in relation to water) and the zones that are not
open may well not be perfectly non-wetting (hydrophobic). This is
why, before proceeding to this second annealing stage, the resin
will first be dissolved, in the acetone for example, for 30 to 40
seconds for example. Preferably, but not necessarily, this
dissolving stage is followed by a rinsing stage, in alcohol for
example.
[0086] Finally, the second annealing stage is performed, at
170.degree. C. for example (according to the material chosen) for 5
minutes, the result of which is to cause any the solvent that may
be present in the hydrophobic material to disappear totally. In
order to obtain a uniform surface and maximum adherence of the
non-wetting material on the substrate, another complementary
annealing process can be effected, at 330.degree. C. for 15 minutes
for example.
[0087] Thus, the method of the invention advantageously allows the
creation of a partially-wetting layer in a non-wetting material.
This result is achieved by the creation of openings in the
non-wetting material, which then become wetting zones, suitable for
chemical or biochemical functionalisation. The zones that are not
open remain perfectly non-wetting, and therefore retain their
enhanced properties of non-wettability which are necessary for the
transportation of drops. In particular, the fact that the layer of
non-wetting material is deposited at the last stage of the method,
in contrast to previous designs, means that this material will not
be subjected to such surface treatment (a technique using a
surface-active agent, or using a plasma-argon).
[0088] The device of the invention therefore includes at least one
layer which is rendered partially wetting by the creation of
wetting openings in a non-wetting layer, as explained previously.
It will be possible to activate and functionalise these wetting
zones chemically (FIG. 8) so as to then react with the manipulated
drop (FIG. 9). Use will therefore be made of the principle for
displacement of the drop as explained previously in order to
activate the zones that are still not functionalised, using a drop
containing an agent that allows functionalisation.
[0089] It can be seen, in particular in FIG. 8 (method of
representation identical to that of the top part of FIGS. 3 and 4,
from above and partial, meaning without the insulating dielectric
and non-wetting layers respectively, between the interdigitated
electrodes and the drop) that a drop containing an agent that
allows the functionalisation 15, starting from electrode 1, moves
to electrode 2 over a functionalisable zone 5, and then arrives at
electrode 3 after having activated and functionalised the zone 5
chemically.
[0090] In FIG. 9 (a method of representation identical to that of
the top part of FIGS. 3 and 4, from above and partial, meaning
without the insulating dielectric and non-wetting layers
respectively, between the interdigitated electrodes and the drop),
it can be seen how a drop 15 moving on the track rests firstly on
electrode 1 and then passes on to electrode 2, above which lies the
functionalised zone 5, and arrives, changed, at electrode 3, after
reaction with the functionalised zone.
[0091] FIG. 10 schematically represents an implementation variant
of the system according to the invention. The system includes one
or more means 1 for preparation of the liquid sample to be
analysed, one or more devices 2 for handling of drops according to
the invention and as explained previously, and one or more means 3
for analysis on exiting. The preparation means 1 can include one or
more loading reservoirs or docks for example. The analysis means 3
can be a mass spectrometer, a fluorescence detector or a UV light
detector for example. The device 2 according to the invention, at
the heart of this system, is coupled upstream with the preparation
means (s) 1, and downstream with the analysis means (s) 3.
[0092] The system according to the invention can be thus possibly
be integrated into a microsystem that itself includes one or more
laboratory operations usually effected manually. Such a system is
known as a microlaboratory.
[0093] Two examples of functionalisation will now be described, on
the basis of an example of implementation of the device of the
invention that includes a substrate in Pyrex.RTM., conducting
interdigitated electrodes in nickel with a thickness of about one
hundred nanometres, a layer of about one micrometre of SU8 resin
deposited by centrifuging, and a dielectric insulating layer.
Finally, the device includes a hydrophobic layer in
tetrafluoroethylene polymer, also deposited by centrifuging, on the
resin layer previously mentioned.
Example of an Affinity Reactor:
[0094] The zones not covered by the hydrophobic layer will undergo
a surface treatment that is intended to convert them into a
reactive surface, such as a Streptavidine grafted NH2 support.
[0095] Thus, with such a device, including such functionalised
zones, a drop of liquid containing proteins for example, and moving
in the path of electrodes over a functionalised zone, will find
that its molecules of interest (certain proteins such as biotine
for example) with an affinity for the surfaces previously grafted
during the functionalisation, fix onto these surfaces. When the
chemical reaction has ended, the drop continues on its path in the
device. In what follows, the passage of a special mixture (a
denaturing buffer mixture for example) in these zones, allows the
molecules of interest to be liberated (by destruction of the
non-covalent interactions for example) and draws them along with
it. Such a device is therefore used to isolate and separate
molecules of interest.
Example of a Digestion Reactor:
[0096] In the device, the zones not covered by the hydrophobic
layer will undergo a surface treatment with the aim of converting
them into reactive surfaces, such as a trypsine grafted NH2 support
for example.
[0097] Thus, in such a device with such functionalised zones, a
drop of liquid moving in the path of electrodes is immobilised in a
functionalised zone, and certain molecules of interest (proteins
for example) will react with the grafted surfaces. The result of
such a reaction will be to cut the molecules (peptides obtained by
tryptidic digestion for example). In what follows, the drop
continues on its path in the device. Such a device therefore allows
the analysis of long chains of molecules for example, by prior
cutting using specific enzymes, with a view to analysis by mass
spectrometry.
[0098] The device, the method, and the system of the invention,
therefore allow implementation of the basic elements of a
microsystem that is intended to move microdroplets from one
functionalised zone to another, in an architecture which lends
itself readily to integration, upstream or downstream, with other
complementary functions. It is therefore possible to design
specialised Microsystems that differ from each other only by the
chaining and the nature of the biochemical operations effected.
[0099] All of the above description is given by way of an example,
and does not limit the invention in any way. In particular, the
choice of a material in tetrafluoroethylene polymers for the
non-wetting or partially wetting layer does not limit the
invention. A tetrafluoroethylene polymer is a suitable choice in
the sense that it is actually non-wetting, in particular, but not
only, in relation to water, and therefore hydrophobic. More
generally, one is always looking for a non-wetting material which
is biocompatible (does not adsorb any of the material transported,
does not mix with the material transported, does not provoke
chemical reactions, and does not leech material) . It must
therefore be neutral in the light of the preceding explanations,
and also display a homogeneity of its surface properties.
[0100] Likewise, the choice of silicon or Pyrex.RTM. for the
substrate does not in any way limit the invention. This is also the
case for the choice of a positive or negative resin in the context
of the method for the manufacture of the device of the invention.
It will also be noted, still in the context of the method for the
manufacture of the device of the invention, that the temperatures
and times of the annealing stages of the method do not limit the
invention, and are essentially a function of the non-wetting
material chosen. In addition, the use of acetone for dissolving and
of alcohol for rinsing, does not limit the invention. Any other
product suitable for dissolving and rinsing can be used.
[0101] Furthermore, the examples of displacement in a given
direction, mentioned in this description, do not limit the
invention. It is naturally possible to envisage a displacement
matrix that allows the drop to be moved anywhere on the track. The
displacement options depend essentially on the geometric layout of
the electrodes. A matrix of electrodes can in fact be used to
achieve a displacement of the matricial type. Also, the shape of
the electrodes in the examples of this description does not in any
way limit the invention. Any other shape allowing interdigitation
of the electrodes will be suitable.
[0102] In addition, the list of the examples of means for the
preparation of the displacement device upstream, in an integrated
system such as the system of the invention, is naturally not
exhaustive, and therefore does not limit the invention. This also
applies to the list of means for analysis of the displacement
device downstream.
[0103] Finally, the examples of functionalisation of the wetting
zones of the partially-wetting layer, and the examples of treatment
of the drop by these functionalised zones, given in this
description, do not limit the invention. Generally, in fact one is
interested in the separation, the sorting or the cutting of
molecules, whatever they may be. Other handlings by chemical and/or
biochemical reactions can also be envisaged.
* * * * *