U.S. patent application number 11/080024 was filed with the patent office on 2006-09-21 for avoidance of bouncing and splashing in droplet-based fluid transport.
Invention is credited to Stephen J. Hinkson, Richard G. Stearns.
Application Number | 20060210443 11/080024 |
Document ID | / |
Family ID | 36592893 |
Filed Date | 2006-09-21 |
United States Patent
Application |
20060210443 |
Kind Code |
A1 |
Stearns; Richard G. ; et
al. |
September 21, 2006 |
Avoidance of bouncing and splashing in droplet-based fluid
transport
Abstract
A system for fluid transport is provided where a quantity of
fluid is held in a reservoir. A droplet generator is employed to
generate droplets from the fluid, for example a nozzle-based system
or a nozzleless system such as an acoustic ejection system. A
generated droplet has a trajectory whereby it arrives at a target.
A circuit is used to modify one or more characteristics of the
generated droplet in a way which increases the likelihood that the
droplet will not splash or bounce when it arrives at the target.
The circuit may in different embodiments control the speed of the
droplet or the Weber number of the droplet. The circuit may create
an electric field in an area of space where the droplet passes. The
circuit may charge the droplet by causing it to contact ions.
Inventors: |
Stearns; Richard G.;
(Felton, CA) ; Hinkson; Stephen J.; (Berkeley,
CA) |
Correspondence
Address: |
MINTZ, LEVIN, COHN, FERRIS, GLOVSKY AND POPEO, P.C
1400 PAGE MILL ROAD
PALO ALTO
CA
94304-1124
US
|
Family ID: |
36592893 |
Appl. No.: |
11/080024 |
Filed: |
March 14, 2005 |
Current U.S.
Class: |
422/400 |
Current CPC
Class: |
G01N 35/1074 20130101;
B05B 17/06 20130101; Y10T 436/2575 20150115; G01N 2035/1034
20130101; B01L 3/0268 20130101; B05B 5/005 20130101; B01L 2400/0415
20130101; B01L 2300/0819 20130101; B01L 2300/0829 20130101; G01N
35/1009 20130101; B01L 2400/027 20130101; B01L 2400/0487 20130101;
B01L 2400/0436 20130101 |
Class at
Publication: |
422/100 |
International
Class: |
B01L 3/02 20060101
B01L003/02 |
Claims
1. A system for fluid transport comprising: a reservoir containing
a quantity of fluid, a droplet generator for generating a droplet
from the fluid in the reservoir, a controller for controlling the
operation of the droplet generator, a target where the generated
droplet arrives, a circuit for controlling one or more
characteristics of the generated droplet which increases the
likelihood that the droplet will not splash or bounce on arriving
at the target.
2. The system of claim 1, wherein the circuit for controlling
generates an electric field in a zone of space through which the
droplet passes.
3. The system of claim 2, wherein the generated electric field is
non-negligible in a zone proximate to a free surface of the fluid
in the reservoir.
4. The system of claim 3, wherein the generated electric field has
a magnitude between 1000 and 100,000 V/m in a zone of space
proximate to a free surface of the fluid in the reservoir.
5. The system of claim 2, wherein the generated electric field is
produced with the assistance of an electrode proximate to the
target.
6. The system of claim 2, wherein the generated electric field
causes the generated droplet to be electrically charged.
7. The system of claim 2, wherein the circuit for controlling
comprises an electrode which is held at a predetermined voltage for
a predetermined period of time.
8. The system of claim 2, wherein the generated electric field is
time varying.
9. The system of claim 2, wherein the generated electric field does
not cause the generated droplet to deviate substantially from a
path the droplet would travel in the absence of the generated
electric field.
10. The system of claim 9, wherein the path comprises the path
traced by the generated droplet beginning with ejection and ending
with impact of the droplet on the target.
11. The system of claim 2, wherein the circuit for controlling
comprises an input allowing external logic to control a magnitude
of the electric field.
12. The system of claim 2, wherein the circuit for controlling
comprises an input allowing external logic to switch a magnitude of
the electric field between a predetermined value and zero.
13. The system of claim 2, wherein the circuit for controlling
comprises a charging device which charges some or all of the target
or an object located in the vicinity of the target.
14. The system of claim 1, wherein the circuit for controlling
causes the velocity of the droplet in the vicinity of the target to
lie within a predetermined range of velocities.
15. The system of claim 14, wherein the predetermined range of
velocities lies within the range of 1.0 to 2.5 m/s.
16. The system of claim 1, wherein the circuit for controlling
causes the Weber number of the droplet in the vicinity of the
target to lie within a predetermined range.
17. The system of claim 1, wherein the droplets travel in a
direction which is at an angle of 90 degrees to 180 degrees to the
direction of the earth's gravitational field as the droplets arrive
at the target.
18. The system of claim 1, wherein the droplet generator is
nozzleless.
19. The system of claim 1, wherein the droplet generator does not
make contact with the fluid in order to generate a droplet.
20. The system of claim 1, wherein the droplet generator comprises
an acoustic ejection system.
21. The system of claim 1, wherein the circuit for controlling
increases the likelihood of coalescence of the droplet with a
second fluid.
22. The system of claim 1, wherein the circuit increases the
probability that the droplet will not splash or bounce when there
is fluid present at the target which has a different composition
from the fluid in the droplet.
23. The system of claim 1, wherein the circuit increases the
probability that the droplet will not splash or bounce when there
is fluid present at the target which has a volume less than twice
that of the droplet.
24. The system of claim 1, wherein the circuit increases the
probability that the droplet will not splash or bounce when there
is fluid present at the target which has a volume more than 100
times that of the droplet.
25. The system of claim 1, wherein the volume of the droplet is
less than 100 nL.
26. The system of claim 25, wherein the volume of the droplet is
less than 5 nL.
27. The system of claim 1, wherein the reservoir forms part of a
well plate.
28. The system of claim 1, comprising a second reservoir, wherein
the droplet generator can generate a droplet from fluid in the
second reservoir.
29. The system of claim 28, comprising a mechanism for moving the
droplet generator to facilitate its generation of a droplet from
fluid in the second reservoir.
30. The system of claim 28, wherein the circuit for controlling
increases the likelihood that a droplet generated from fluid in the
second reservoir arrives at a second target without splashing or
bouncing.
31. The system of claim 28, wherein the circuit for controlling
increases the likelihood that a droplet generated by the second
reservoir coalesces with a second fluid.
32. A method for fluid transport comprising the steps of:
generating a droplet from a quantity of fluid in a reservoir,
controlling a trajectory of the droplet in such a way that it
arrives at a target, controlling one or more characteristics of the
generated droplet so as to increase the likelihood that the droplet
will not splash or bounce on arriving at the target.
33. The method of claim 32, wherein the step of controlling one or
more characteristics comprises generating an electric field in a
zone of space through which the droplet passes.
34. The method of claim 33, wherein the generated electric field is
non-negligible in a zone proximate to a free surface of the fluid
in the reservoir.
35. The method of claim 34, wherein the generated electric field
has a magnitude between 1000 and 100,000 V/m in a zone of space
proximate to a free surface of the fluid in the reservoir.
36. The method of claim 33, wherein the generated electric field is
produced with the assistance of an electrode proximate to the
target.
37. The method of claim 33, wherein the generated electric field
causes the generated droplet to be electrically charged.
38. The method of claim 33, wherein the step of generating an
electric field comprises holding an electrode at a predetermined
voltage for a predetermined period of time.
39. The method of claim 33, wherein the generated electric field is
time varying.
40. The method of claim 33, wherein the generated electric field
does not cause the generated droplet to deviate substantially from
a path the droplet would travel in the absence of the generated
electric field.
41. The method of claim 40, wherein the path comprises the path
traced by the generated droplet beginning with ejection and ending
with impact of the droplet on the target.
42. The method of claim 33, wherein the step of generating an
electric field comprises setting the magnitude of the electric
field in response to an external input.
43. The method of claim 33, wherein the step of generating an
electric field comprises switching a magnitude of the electric
field between a predetermined value and zero.
44. The method of claim 32, wherein the step of controlling one or
more characteristics comprises causing the velocity of the droplet
in the vicinity of the target to lie within a predetermined range
of velocities.
45. The method of claim 44, wherein the predetermined range of
velocities lies within the range of 1.0 to 2.5 m/s.
46. The method of claim 32, wherein the step of controlling one or
more characteristics comprises causing the Weber number of the
droplet in the vicinity of the target to lie within a predetermined
range.
47. The method of claim 32, wherein the droplets travel in a
direction which is at an angle of 90 degrees to 180 degrees to the
direction of the earth's gravitational field as the droplets arrive
at the target.
48. The method of claim 32, wherein the step of generating a
droplet employs a nozzleless droplet generator.
49. The method of claim 32, wherein the step of generating a
droplet does not cause an external object to contact the fluid.
50. The method of claim 32, wherein the step of generating a
droplet comprises the step of directing focused acoustic energy at
a free surface of the quantity of fluid.
51. The method of claim 32, wherein the step of controlling one or
more characteristics increases the likelihood of coalescence of the
droplet with a second fluid.
52. The method of claim 32, wherein the reservoir forms part of a
well plate.
53. The method of claim 32, comprising the step of generating a
droplet of fluid from a second reservoir.
54. The method of claim 53, wherein the step of generating a
droplet from a second reservoir comprises the step of moving a
droplet generator.
55. The method of claim 53, wherein the step of controlling one or
more characteristics increases the likelihood that a droplet
generated from fluid in the second reservoir arrives at a second
target without splashing or bouncing.
56. The method of claim 53, wherein the step of controlling one or
more characteristics increases the likelihood that a droplet
generated by the second reservoir coalesces with a second
fluid.
57. The method of claim 32, wherein the step of controlling one or
more characteristics increases the probability that the droplet
will not splash or bounce when there is fluid-present at the target
which has a different composition from the fluid in the
droplet.
58. The method of claim 32, wherein the step of controlling one or
more characteristics increases the probability that the droplet
will not splash or bounce when there is fluid present at the target
which has a volume less than twice that of the droplet.
59. The method of claim 32, wherein the step of controlling one or
more characteristics increases the probability that the droplet
will not splash or bounce when there is fluid present at the target
which has a volume more than 100 times that of the droplet.
60. The method of claim 32, wherein there is fluid present at the
target which has a different composition from the fluid in the
droplet and the droplet contacts the fluid present at the
target.
61. The method of claim 32, wherein there is fluid present at the
target which has a volume less than twice that of the droplet and
the droplet contacts the fluid present at the target.
62. The method of claim 32, wherein there is fluid present at the
target which has a volume more than 100 times that of the droplet
and the droplet contacts the fluid present at the target.
63. The method of claim 32, wherein there is fluid present at the
target which has a viscosity different from that of the droplet and
the droplet contacts the fluid present at the target.
64. The method of claim 63, wherein there is fluid present at the
target which has a viscosity differing by at least 50% from that of
the droplet and the droplet contacts the fluid present at the
target.
65. The method of claim 64, wherein there is fluid present at the
target which has a viscosity at least three times that of the
droplet and the droplet contacts the fluid present at the
target.
66. The method of claim 63, wherein there is fluid present at the
target which has a viscosity no more than one-third that of the
droplet and the droplet contacts the fluid present at the
target.
67. The method of claim 33, wherein the step of generating an
electric field comprises imparting an electrostatic charge to all
or some part of the target or to an object in the vicinity of the
target.
68. The method of claim 32, wherein the step of controlling one or
more characteristics comprises imparting an electrostatic charge to
the droplet.
69. The method of claim 65, wherein the step of imparting an
electrostatic charge to the droplet comprises contacting the
droplet with ions.
Description
TECHNICAL FIELD
[0001] This invention relates generally to systems and methods for
transporting fluids, and specifically to systems and methods for
transporting fluids as droplets.
BACKGROUND
[0002] There exists a need in pharmaceutical, biotechnological,
medical, and other industries to be able to quickly screen,
identify, analyze, and/or process large numbers or varieties of
fluids. As a result, much attention has been focused on developing
efficient, precise, and accurate fluid handling methods. For
example, automated robotic systems have been used in combination
with precise registration technologies to dispense reagents through
automated pick-and-place ("suck-and-spit") fluid handling systems.
Similarly, some efforts have been directed to adapting printing
technologies, particularly inkjet printing technologies, to form
biomolecular arrays. For example, U.S. Pat. No. 6,015,880 to
Baldeschwieler et al. is directed to array preparation using
multistep in situ synthesis. Such synthesis may involve using
inkjet technology to dispense reagent-containing droplets to a
locus on a surface chemically prepared to permit covalent
attachment of the reagent.
[0003] There are tradeoffs in the choice of a fluid transport
system. For example, most fluid handling systems presently in use
require that contact be established between the fluid to be
transferred and an associated solid surface on the transferring
device. Such contact typically results in surface wetting that
causes unavoidable fluid waste, a notable drawback when the fluid
to be transferred is rare and/or expensive. When fluid transport
systems are constructed using networks of tubing or other fluid
transporting conduits, air bubbles can be entrapped or particulates
may become lodged in the networks. Nozzles of ordinary inkjet
printheads are also subject to clogging, especially when used to
eject a macromolecule-containing fluid at elevated temperatures, a
situation commonly associated with such technologies. As a result,
ordinary fluid transport technologies may produce improperly sized
or misdirected droplets.
[0004] A number of patents have described the use of focused
acoustic radiation to dispense fluids such as inks and reagents.
For example, U.S. Pat. No. 4,308,547 to Lovelady et al. describes a
liquid drop emitter that utilizes acoustic principles to eject
droplets from a body of liquid onto a moving document to result in
the formation of characters or barcodes thereon. A nozzleless
inkjet printing apparatus is used such that controlled drops of ink
are propelled by an acoustical force produced by a curved
transducer at or below the surface of the ink. Similarly, U.S.
Patent Application Publication No. 20020037579 to Ellson et al.
describes a device for acoustically ejecting a plurality of fluid
droplets toward discrete sites on a substrate surface for
deposition thereon. U.S. Patent Application Publication No.
20020094582 to Williams describes technologies that employ focused
acoustic technology as well. In contrast to inkjet printing
devices, focused acoustic radiation may be used to effect
nozzleless fluid ejection, and devices using focused acoustic
radiation are not generally subject to clogging and the
disadvantages associated therewith, e.g., misdirected fluid or
improperly sized droplets.
[0005] Since fluids used in pharmaceutical, biotechnological, and
other scientific industries may be rare and/or expensive,
techniques capable of handling small volumes of fluids provide
readily apparent advantages over those requiring relatively larger
volumes. Typically, fluids for use in combinatorial methods are
provided as a collection or library of organic and/or biological
compounds. In many instances, well plates are used to store a large
number of fluids for screening and/or processing. Well plates are
typically of single piece construction and comprise a plurality of
identical wells, wherein each well is adapted to contain a small
volume of fluid. Such well plates are commercially available in
standardized sizes and may contain, for example, 96, 384, 1536, or
3456 wells per well plate.
[0006] Transport of fluid droplets may be directed at an existing
volume of fluid. For example, in any fluid transport system that
employs discrete droplets, it may be desirable to use a number of
smaller droplets to transport the fluid rather than a single larger
droplet. Each droplet after the first will potentially impact an
existing volume of fluid.
[0007] When a fluid droplet is directed at an existing volume of
fluid, it is often desirable that the droplet coalesce with the
existing volume. Instead of coalescing, a droplet may bounce or
splash, which is often undesirable. Bouncing and splashing may also
be undesirable when the droplets are directed at a solid target.
For example, the target may be a well plate in which the droplet is
supposed to be placed entirely in an identified individual well in
accordance with the transfer protocol being employed, whereas
splashing might cause a portion of the fluid in the droplet to fall
into a different well instead.
[0008] Bouncing of droplets when they encounter a solid or an
existing volume of fluid has been observed for many years. It is
believed that the phenomenon involves not simply the droplet and
the solid or volume of fluid but also a cushion of air between the
droplet and the solid or volume. Precise predictions of when
droplet bouncing and splashing will occur based on conventional
fluid parameters such as viscosity and surface tension are often
not within the capabilities of computational fluid mechanics, so
that empirical investigation is a preferred method of analyzing
questions which relate to droplet bouncing and splashing. A summary
of certain empirical investigations is found in M. Orme,
"Experiments on Droplet Collisions, Bounce, Coalescence and
Disruption," Progress in Energy and Combustion Science, vol. 23,
pp. 65-79, 1997, which contains a number of references to the
literature.
SUMMARY OF THE INVENTION
[0009] The invention is in general a system for fluid transport. A
quantity of fluid is held in a reservoir. A droplet generator is
employed to generate droplets from the fluid, for example a
nozzle-based system or a nozzleless system such as an acoustic
ejection system. The droplet generator may or may not make contact
with the fluid in order to generate the droplet. The droplet
generator may be set up to move from one reservoir to another in
order to be able to eject from more than one reservoir. In a common
arrangement, a number of reservoirs form part of an integral
structure, e.g., a well plate, and the structure is moved by
suitable mechanical or electromechanical systems, potentially under
computer control, into a suitable position with respect to the
droplet generator.
[0010] The droplet generator is controlled by a controller,
preferably electronic and connectable to computers through some
communications system. The droplets after generation arrive at a
target, which may be for example a well plate, a porous or
non-porous surface, a substrate, or a structure which is to be
coated by the droplets. A circuit controls one or more
characteristics of the generated droplet, increasing the likelihood
that the droplet will not splash or bounce on arriving at the
target. This circuit may be controlled by the controller or may
form part of the controller. The target may already have a quantity
of fluid, and the droplet may coalesce with that quantity of
fluid.
[0011] In certain preferred embodiments, the circuit for
controlling one or more characteristics may control the speed of
the droplet as it arrives at the target. The speed may be
controlled in preferred ranges. In another embodiment, the circuit
for controlling may control the Weber number of the droplet. In
doing so, the circuit may control for example the diameter of the
droplet or its velocity or both. In another preferred embodiment,
the circuit may provide an electric field in an area of space
through which the droplet passes. This electric field may result in
polarization or charging of the droplet. In another preferred
embodiment, the circuit may charge the droplet by causing it to
contact ions.
[0012] The invention also encompasses methods for the transport of
fluids. In these methods, a droplet is generated from a quantity of
fluid in a reservoir. A trajectory of the droplet is controlled in
such a way that it arrives at a target. One or more characteristics
of the generated droplet are controlled so as to increase the
likelihood that the droplet will not splash or bounce on arriving
at the target.
BRIEF DESCRIPTION OF THE DRAWINGS
[0013] The invention is described in detail below with reference to
the following drawings, wherein like reference numerals indicate a
corresponding structure throughout the several views.
[0014] FIGS. 1A and 1B, collectively referred to as FIG. 1,
schematically illustrate in simplified cross-sectional view the
operation of a focused acoustic ejection device in the preparation
of a plurality of features on a substrate surface. FIG. 1A shows
the acoustic ejector acoustically coupled to a first reservoir and
having been activated in order to eject a first droplet of fluid
from within the reservoir toward a particular site on a substrate
surface. FIG. 1B shows the acoustic ejector acoustically coupled to
a second reservoir and having been activated to eject a second
droplet of fluid from within the second reservoir.
[0015] FIG. 2 illustrates in cross-sectional schematic view the
ejection of droplets of fluid from a volume of fluid on a substrate
surface into an inlet opening disposed on a terminus of a
capillary.
[0016] FIG. 3 depicts the approach of a droplet to a target with an
electric field in the direction of droplet approach.
[0017] FIG. 4 depicts an exemplary circuit of the invention.
[0018] FIGS. 5A-5C show the speed control achievable through
control of the acoustic energy in an acoustic ejection system.
[0019] FIGS. 6A-6C show the effect of droplet speed on the
likelihood of droplet coalescence for 70% DMSO/30% water.
DETAILED DESCRIPTION OF THE INVENTION
[0020] Before describing the present invention in detail, it is to
be understood that this invention is not limited to specific
fluids, biomolecules, or device structures, as such may vary. It is
also to be understood that the terminology used herein is for the
purpose of describing particular embodiments only, and is not
intended to be limiting.
[0021] It must be noted that, as used in this specification and the
appended claims, the singular forms "a," "an," and "the" include
both singular and plural referents unless the context clearly
dictates otherwise. Thus, for example, reference to "a reservoir"
includes a plurality of reservoirs as well as a single reservoir,
reference to "a droplet" includes a plurality of droplets as well
as single droplet, and the like.
[0022] In describing and claiming the present invention, the
following terminology will be used in accordance with the
definitions set out below.
[0023] The terms "acoustic coupling" and "acoustically coupled" as
used herein refer to a state wherein an object is placed in direct
or indirect contact with another object so as to allow acoustic
radiation to be transferred between the objects without substantial
loss of acoustic energy. When two items are indirectly acoustically
coupled, an "acoustic coupling medium" is needed to provide an
intermediary through which acoustic radiation may be transmitted.
Thus, an ejector may be acoustically coupled to a fluid, e.g., by
immersing the ejector in the fluid or by interposing an acoustic
coupling medium between the ejector and the fluid, in order to
transfer acoustic radiation generated by the ejector through the
acoustic coupling medium and into the fluid.
[0024] The term "array" as used herein refers to a two-dimensional
arrangement of features, such as an arrangement of reservoirs
(e.g., wells in a well plate) or an arrangement of different
moieties, including ionic, metallic, or covalent crystalline, e.g.,
molecular crystalline, composite, ceramic, vitreous, amorphous,
fluidic, or molecular materials on a substrate surface (as in an
oligonucleotide or peptidic array). Arrays are generally comprised
of regular features that are ordered, as in, for example, a
rectilinear grid, parallel stripes, spirals, and the like, but
non-ordered arrays may be advantageously used as well. In
particular, the term "rectilinear array" as used herein refers to
an array that has rows and columns of features wherein the rows and
columns typically, but not necessarily, intersect each other at a
ninety-degree angle. An array is distinguished from the more
general term "pattern" in that patterns do not necessarily contain
regular and ordered features. Arrays typically but do not
necessarily comprise at least about 4 to about 10,000,000 features,
generally in the range of about 4 to about 1,000,000 features.
[0025] The terms "biomolecule" and "biological molecule" are used
interchangeably herein to refer to any organic molecule that is,
was, or can be a part of a living organism, regardless of whether
the molecule is naturally occurring, recombinantly produced, or
chemically synthesized in whole or in part. The terms encompass,
for example, nucleotides, amino acids, and monosaccharides, as well
as oligomeric and polymeric species, such as oligonucleotides and
polynucleotides; peptidic molecules, such as oligopeptides,
polypeptides, and proteins; saccharides, such as disaccharides,
oligosaccharides, polysaccharides, and mucopolysaccharides or
peptidoglycans (peptido-polysaccharides); and the like. The terms
also encompass ribosomes, enzyme cofactors, pharmacologically
active agents, and the like. Additional information relating to the
term "biomolecule" can be found in U.S. Patent Application
Publication No. 20020037579 to Ellson et al.
[0026] The term "capillary" is used herein to refer to a conduit
having a bore of small dimension. Typically, capillaries for
electrophoresis that are free standing tubes have an inner diameter
in the range of about 50 to about 250 .mu.m. Capillaries with
extremely small bores integrated to other devices, such as openings
for loading microchannels of microfluidic devices, can be as small
as 1 .mu.m, but in general these capillary openings are in the
range of about 10 to about 100 .mu.M. In the context of delivery to
a mass analyzer in electrospray-type mass spectrometry, the inner
diameter of capillaries may range from about 0.1 to about 3 mm and
preferably from about 0.5 to about 1 mm. In some instances, a
capillary can represent a portion of a microfluidic device. In such
instances, the capillary may be an integral or affixed (permanently
or detachably) portion of the microfluidic device.
[0027] The term "fluid" as used herein refers to matter that is
nonsolid, or at least partially gaseous and/or liquid, but not
entirely gaseous. A fluid may contain a solid that is minimally,
partially, or fully solvated, dispersed, or suspended. Examples of
fluids include, without limitation, aqueous liquids (including
water per se and salt water) and nonaqueous liquids such as organic
solvents and the like. As used herein, the term "fluid" is not
synonymous with the term "ink" in that an ink must contain a
colorant and may not be gaseous.
[0028] The terms "focusing means" and "acoustic focusing means"
refer to a means for causing acoustic waves to converge at a focal
point, either by a device separate from the acoustic energy source
that acts like an optical lens, or by the spatial arrangement of
acoustic energy sources to effect convergence of acoustic energy at
a focal point by constructive and destructive interference. A
focusing means may be as simple as a solid member having a curved
surface, or it may include complex structures such as those found
in Fresnel lenses, which employ diffraction in order to direct
acoustic radiation. Suitable focusing means also include phased
array methods as are known in the art and described, for example,
in U.S. Pat. No. 5,798,779 to Nakayasu et al. and by Amemiya et al.
(1997) Proceedings of the 1997 IS&T NIP 13 International
Conference on Digital Printing Technologies, pp. 698-702.
Additional information regarding acoustic focusing is provided in
U.S. patent application Ser. No. 10/066,546, entitled "Acoustic
Sample Introduction for Analysis and/or Processing," filed Jan. 30,
2002, inventors Ellson and Mutz.
[0029] The terms "library" and "combinatorial library" are used
interchangeably herein to refer to a plurality of chemical or
biological moieties arranged in a pattern or an array such that the
moieties are individually addressable. In some instances, the
plurality of chemical or biological moieties is present on the
surface of a substrate, and in other instances the plurality of
moieties represents the contents of a plurality of reservoirs.
Preferably, but not necessarily, each moiety is different from each
of the other moieties. The moieties may be, for example, peptidic
molecules and/or oligonucleotides.
[0030] The "limiting dimension" of an opening refers herein to the
theoretical maximum diameter of a sphere that can pass through an
opening without deformation. For example, the limiting dimension of
a circular opening is the diameter of the opening. As another
example, the limiting dimension of a rectangular opening is the
length of the shorter side of the rectangular opening. The opening
may be present on any solid body including, but not limited to,
sample vessels, substrates, capillaries, microfluidic devices, and
ionization chambers. Depending on the purpose of the opening, the
opening may represent an inlet and/or an outlet.
[0031] The term "moiety" refers to any particular composition of
matter, e.g., a molecular fragment, an intact molecule (including a
monomeric molecule, an oligomeric molecule, or a polymer), or a
mixture of materials (for example, an alloy or a laminate).
[0032] The term "near," when used to refer to the distance from the
focal point of the focused acoustic radiation to the surface of the
fluid from which a droplet is to be ejected, indicates that the
distance should be such that the focused acoustic radiation
directed into the fluid results in droplet ejection from the fluid
surface; one of ordinary skill in the art will be able to select an
appropriate distance for any given fluid using straightforward and
routine experimentation. Generally, however, a suitable distance
between the focal point of the acoustic radiation and the fluid
surface is in the range of about 1 to about 15 times the wavelength
of the speed of sound in the fluid, more typically in the range of
about 1 to about 10 times that wavelength, preferably in the range
of about 1 to about 5 times that wavelength.
[0033] "Optional" or "optionally" means that the subsequently
described circumstance may or may not occur, so that the
description includes instances where the circumstance occurs and
instances where it does not.
[0034] The term "radiation" is used in its ordinary sense and
refers to emission and propagation of energy in the form of a
waveform disturbance traveling through a medium such that energy is
transferred from one particle of the medium to another, generally
without causing any permanent displacement of the medium itself.
Thus, radiation may refer, for example, to electromagnetic
waveforms as well as acoustic vibrations.
[0035] Accordingly, the terms "acoustic radiation" and "acoustic
energy" are used interchangeably herein and refer to the emission
and propagation of energy in the form of sound waves. As with other
waveforms, acoustic radiation may be focused using a focusing
means, as discussed below. Although acoustic radiation may have a
single frequency and associated wavelength, acoustic radiation may
take a form, e.g. a linear chirp, that includes a plurality of
frequencies. Thus, the term "characteristic wavelength" is used to
describe the mean wavelength of acoustic radiation having a
plurality of frequencies.
[0036] The term "reservoir" as used herein refers to a receptacle
or chamber for containing a fluid. In some instances, a fluid
contained in a reservoir will have a free surface, e.g., a surface
that allows acoustic radiation to be reflected therefrom or a
surface from which a droplet may be acoustically ejected. A
reservoir may also be a locus on a substrate surface within which a
fluid is constrained.
[0037] The term "substrate" as used herein refers to any material
having a surface onto which one or more fluids may be deposited.
The substrate may be constructed in any of a number of forms
including, for example, wafers, slides, well plates, or membranes.
In addition, the substrate may be porous or nonporous as required
for deposition of a particular fluid. Suitable substrate materials
include, but are not limited to, supports that are typically used
for solid phase chemical synthesis, such as polymeric materials
(e.g., polystyrene, polyvinyl acetate, polyvinyl chloride,
polyvinyl pyrrolidone, polyacrylonitrile, polyacrylamide,
polymethyl methacrylate, polytetrafluoroethylene, polyethylene,
polypropylene, polyvinylidene fluoride, polycarbonate, and
divinylbenzene styrene-based polymers), agarose (e.g.,
Sepharose.RTM.), dextran (e.g., Sephadex.RTM.), cellulosic polymers
and other polysaccharides, silica and silica-based materials, glass
(particularly controlled pore glass, or "CPG") and functionalized
glasses, ceramics, and such substrates treated with surface
coatings, e.g., with microporous polymers (particularly cellulosic
polymers such as nitrocellulose), microporous metallic compounds
(particularly microporous aluminum), antibody-binding proteins
(available from Pierce Chemical Co., Rockford Ill.), bisphenol A
polycarbonate, or the like. Additional information relating to the
term "substrate" can be found in U.S. Patent Application
Publication No. 200200377579 to Ellson et al.
[0038] The term "substantially" as in, for example, the phrase
"substantially deviate from a predetermined volume," refers to a
volume that does not deviate by more than about 25%, preferably
10%, more preferably 5%, and most preferably at most 2%, from the
predetermined volume. Other uses of the term "substantially"
involve an analogous definition.
[0039] The term "sample vessel" as used herein refers to any hollow
or concave receptacle having a structure that allows for sample
processing and/or analysis. Thus, a sample vessel has an inlet
opening through which sample may be introduced and an optional, but
preferred, outlet opening through which processed or analyzed
sample may exit.
[0040] The invention may be employed with any type of fluid
dispenser that serves to dispense one or more droplets of fluid
from a reservoir. Any fluid droplet dispensing techniques known in
the art may be used in conjunction with the present invention. For
example, the invention may be used with dispensers such as inkjet
printheads (both thermal and piezoelectric), pipettes, capillaries,
syringes, displacement pumps, rotary pumps, peristaltic pumps,
vacuum devices, flexible or rigid tubing, valves, manifolds,
pressurized gas canisters, and combinations thereof. While
nonacoustic techniques may be used to dispense fluid from the
reservoir, the invention is particularly suited for use with
nozzleless acoustic ejection techniques that employ focused
acoustic radiation generated by acoustic ejectors, such as those
described in U.S. Patent Application Publication No. 20020037579 to
Ellson et al. This publication sets forth that an ejector may be
acoustically coupled to a reservoir containing a fluid in order to
eject a droplet therefrom. In some instances, the reservoir may be
a well of a well plate. Since this device configuration allows
droplets to be ejected from near the base of a well, uncontrolled
electrostatic charge anywhere in the well, e.g., the base or
sidewalls, may have a strong effect influence on the volume and/or
trajectory of such droplets.
[0041] Since acoustic ejection provides a number of advantages over
other fluid dispensing technologies, some embodiments of the
invention employ a device for acoustically ejecting a droplet of
fluid from a reservoir. The device is comprised of a reservoir
adapted to contain a fluid, an ejector for ejecting a droplet from
the reservoir, and a means for positioning the ejector in acoustic
coupling relationship to the reservoir. The ejector comprises an
acoustic radiation generator for generating acoustic radiation and
a focusing means for focusing the acoustic radiation generated by
the generator. As described in U.S. Patent Application Publication
No. 20020037579 to Ellson et al., the acoustic radiation is focused
at a focal point within and sufficiently near the fluid surface in
the reservoir to result in the ejection of droplets therefrom.
Furthermore, a means is provided for reducing any uncontrolled
electrostatic charge on the device or a portion thereof that alters
the volume and/or trajectory of a droplet ejected from the
reservoir. As a result, the volume and/or trajectory of the ejected
droplet do not substantially deviate from a predetermined volume
and/or predetermined trajectory.
[0042] The device may be constructed to include the reservoir as an
integrated or permanently attached component of the device.
However, to provide modularity and interchangeability of
components, it is preferred that the device be constructed with a
removable reservoir. Optionally, a plurality of reservoirs many be
provided. Generally, the reservoirs are arranged in a pattern or an
array to provide each reservoir with individual systematic
addressability. In addition, while each of the reservoirs may be
provided as a discrete or stand-alone item, in circumstances that
require a large number of reservoirs, it is preferred that the
reservoirs be attached to each other or represent integrated
portions of a single reservoir unit. For example, the reservoirs
may represent individual wells in a well plate.
[0043] Many well plates suitable for use with the device are
commercially available and may contain, for example, 96, 384, 1536,
or 3456 wells per well plate, having a full skirt, half skirt, or
no skirt. The wells of such well plates typically form rectilinear
arrays. Manufacturers of suitable well plates for use in the
employed device include Corning, Inc. (Corning, N.Y.) and Greiner
America, Inc. (Lake Mary, Fla.). However, the availability of such
commercially available well plates does not preclude the
manufacture and use of custom-made well plates containing at least
about 10,000 wells, or as many as 100,000 to 500,000 wells, or
more. The wells of such custom-made well plates may form
rectilinear or other types of arrays. As well plates have become
commonly used laboratory items, the Society for Biomolecular
Screening (Danbury, Conn.) has formed the Microplate Standards
Development Committee to recommend and maintain standards to
facilitate the automated processing of small volume well plates on
behalf of and for acceptance by the American National Standards
Institute.
[0044] Furthermore, the material used in the construction of
reservoirs must be compatible with the fluids contained therein.
Thus, if it is intended that the reservoirs or wells contain an
organic solvent such as acetonitrile, polymers that dissolve or
swell in acetonitrile would be unsuitable for use in forming the
reservoirs or well plates. Similarly, reservoirs or wells intended
to contain DMSO must be compatible with DMSO. For water-based
fluids, a number of materials are suitable for the construction of
reservoirs and include, but are not limited to, ceramics such as
silicon oxide and aluminum oxide, metals such as stainless steel
and platinum, and polymers such as polyester and
polytetrafluoroethylene. For fluids that are photosensitive, the
reservoirs may be constructed from an optically opaque material
that has sufficient acoustic transparency for substantially
unimpaired functioning of the device. Thus, the reservoir may be
adapted to contain any type of fluid, metallic or nonmetallic,
organic or inorganic.
[0045] When a plurality of reservoirs is employed, the acoustic
radiation generator may have to be aligned with each reservoir
during operation, discussed infra. In order to reduce the amount of
movement and time needed to align the generator successively with
each reservoir, it is preferable that the center of each reservoir
be located not more than about 1 centimeter, more preferably not
more than about 1.5 millimeters, still more preferably not more
than about 1 millimeter and optimally not more than about 0.5
millimeter, from a neighboring reservoir center. These dimensions
tend to limit the size of the reservoirs to a maximum volume. The
reservoirs are constructed to contain typically no more than about
1 mL, preferably no more than about 100 .mu.L, more preferably no
more than about 10 .mu.L, still more preferably no more than about
1 .mu.L, and optimally no more than about 1 nL, of fluid. The
reservoirs may be either completely or partially filled with fluid.
For example, fluid may occupy a volume of about 10 pL to about 100
nL.
[0046] When an array of reservoirs is provided, each reservoir may
be individually, efficiently, and systematically addressed.
Although any type of array may be employed, arrays comprised of
parallel rows of evenly spaced reservoirs are preferred. Typically,
though not necessarily, each row contains the same number of
reservoirs. Optimally, rectilinear arrays comprising X rows and Y
columns of reservoirs are employed with the invention, wherein X
and Y are each at least 2. In some instances, X may be greater
than, equal to, or less than Y. In addition, nonrectilinear arrays
as well as other geometries may be employed. For example,
hexagonal, spiral, or other types of arrays may be used. In some
instances, the invention may be employed with irregular patterns of
reservoirs, e.g., droplets randomly located on a flat substrate
surface. In addition, the invention may be used with reservoirs
associated with microfluidic devices.
[0047] Moreover, the invention may be used to transport fluids of
virtually any type and amount desired. The fluid may be aqueous
and/or nonaqueous. Examples of fluids include, but are not limited
to, aqueous fluids including water per se and water-solvated ionic
and non-ionic solutions; organic solvents; lipidic liquids;
suspensions of immiscible fluids; and suspensions or slurries of
solids in liquids. Because the invention is readily adapted for use
with high temperatures, fluids such as liquid metals, ceramic
materials, and glasses may be used, as described in U.S. Patent
Application Publication No. 20020140118. In some instances, the
reservoir may contain a biomolecule, nucleotidic, peptidic, or
otherwise. In addition, the invention may be used in conjunction
with dispensers for dispensing droplets of immiscible fluids, as
described in U.S. Patent Application Publication Nos. 2002037375
and 20020155231, or to dispense droplets containing pharmaceutical
agents, as discussed in U.S. Patent Application Publication No.
20020142049 and U.S. patent application Ser. No. 10/244,128,
entitled "Precipitation of Solid Particles from Droplets Formed
Using Focused Acoustic Energy," filed, Sep. 13, 2002, inventors
Lee, Ellson and Williams.
[0048] Any of a variety of focusing means may be employed to focus
acoustic radiation so as to eject droplets from a reservoir. For
example, one or more curved surfaces may be used to direct acoustic
radiation to a focal point near a fluid surface. One such technique
is described in U.S. Pat. No. 4,308,547 to Lovelady et al. Focusing
means with a curved surface have been incorporated into the
construction of commercially available acoustic transducers such as
those manufactured by Panametrics Inc. (Waltham, Mass.). In
addition, Fresnel lenses are known in the art for directing
acoustic energy at a predetermined focal distance from an object
plane. See, e.g., U.S. Pat. No. 5,041,849 to Quate et al. Fresnel
lenses may have a radial phase profile that diffracts a substantial
portion of acoustic energy into a predetermined diffraction order
at diffraction angles that vary radially with respect to the lens.
The diffraction angles should be selected to focus the acoustic
energy within the diffraction order on a desired object plane. It
should be noted that acoustic focusing means exhibiting a variety
of F-numbers may be employed with the invention. As discussed in
U.S. Pat. No. 6,416,164 to Stearns et al., however, low F-number
focusing places restrictions on the reservoir and fluid level
geometry and provides relatively limited depth of focus, increasing
the sensitivity to the fluid level in the reservoir. Thus, the
focusing means suitable for use with the invention typically
exhibits an F-number of at least about 1. Preferably, the focusing
means exhibits an F-number of at least about 2.
[0049] There are a number of ways to acoustically couple the
ejector to a reservoir and thus to the fluid therein. One such
approach is through direct contact, as is described, for example,
in U.S. Pat. No. 4,308,547 to Lovelady et al., wherein a focusing
means constructed from a hemispherical crystal having segmented
electrodes is submerged in a liquid to be ejected. The
aforementioned patent further discloses that the focusing means may
be positioned at or below the surface of the liquid. However, this
approach for acoustically coupling the focusing means to a fluid is
undesirable when the ejector is used to eject different fluids in a
plurality of containers or reservoirs, as repeated cleaning of the
focusing means would be required in order to avoid
cross-contamination. The cleaning process would necessarily
lengthen the transition time between each droplet ejection event.
In addition, in such a method, fluid would adhere to the ejector as
it is removed from each container, wasting material that may be
costly or rare.
[0050] Thus, a preferred approach is to acoustically couple the
ejector to the reservoir without contacting any portion of the
ejector, e.g., the focusing means, with the fluids to be ejected.
When a plurality of reservoirs is employed, a positioning means is
provided for positioning the ejector in controlled and repeatable
acoustic coupling with each of the fluids in the reservoirs to
eject droplets therefrom without submerging the ejector therein.
This typically involves direct or indirect contact between the
ejector and the external surface of each reservoir. When direct
contact is used in order to acoustically couple the ejector to each
reservoir, it is preferred that the direct contact be wholly
conformal to ensure efficient acoustic energy transfer. That is,
the ejector and the reservoir should have corresponding surfaces
adapted for mating contact. Thus, if acoustic coupling is achieved
between the ejector and reservoir through the focusing means, it is
desirable for the reservoir to have an outside surface that
corresponds to the surface profile of the focusing means. Without
conformal contact, efficiency and accuracy of acoustic energy
transfer may be compromised. In addition, since many focusing means
have a curved surface, the direct contact approach may necessitate
the use of reservoirs having a specially formed inverse
surface.
[0051] When an ejector is placed in indirect contact with a
reservoir, an acoustic coupling medium may be interposed between
the reservoir and ejector. Typically, the acoustic coupling medium
is a fluid. In addition, the acoustic coupling medium is preferably
an acoustically homogeneous material that is substantially free of
material having different acoustic properties than the fluid medium
itself. Furthermore, it is preferred that the acoustic coupling
medium be comprised of a material having acoustic properties that
facilitate the transmission of acoustic radiation without
significant attenuation in acoustic pressure and intensity. Also,
the acoustic impedance of the coupling medium should facilitate the
transfer of energy from the coupling medium into the reservoir. An
aqueous fluid, such as water per se, may be employed as an acoustic
coupling medium. Ionic additives, e.g., salts, may sometimes be
added to the coupling medium to increase the conductivity of the
coupling medium.
[0052] A single ejector is preferred, although an acoustic ejection
system may include a plurality of ejectors. When a single ejector
is employed, the means for positioning the ejector may be adapted
to provide relative motion between the ejector and reservoirs. The
positioning means should allow for the ejector to move from one
reservoir to another quickly and in a controlled manner, thereby
allowing fast and controlled scanning of the reservoirs to effect
droplet ejection therefrom. Thus, various means for positioning the
ejector in acoustic coupling relationship to the reservoir are
generally known in the art and may involve, e.g., devices that
provide movement having one, two, three, four, five, six, or more
degrees of freedom. Accordingly, when rows of reservoirs are
provided, relative motion between the acoustic radiation generator
and the reservoirs may result in displacement of the acoustic
radiation generator in a direction along the rows. Similarly, when
a rectilinear array of reservoirs is provided, the ejector may be
movable in a row-wise direction and/or in a direction perpendicular
to both the rows and columns.
[0053] Current positioning technology allows for the ejector
positioning means to move from one reservoir to another quickly and
in a controlled manner, thereby allowing fast and controlled
ejection of different fluid samples. That is, current commercially
available technology allows the ejector to be moved from one
reservoir to another, with repeatable and controlled acoustic
coupling at each reservoir, in less than about 0.1 second for high
performance positioning means and in less than about 1 second for
ordinary positioning means. A custom designed system will allow the
ejector to be moved from one reservoir to another with repeatable
and controlled acoustic coupling in less than about 0.001
second.
[0054] Acoustic ejection also enables rapid ejection of droplets
from one or more reservoirs, e.g., at a rate of at least about
1,000,000 droplets per minute from the same reservoir, and at a
rate of at least about 100,000 drops per minute from different
reservoirs, assuming that the droplet size does not exceed about 10
.mu.m in diameter. One of ordinary skill in the art will recognize
that the droplet generation rate is a function of drop size,
viscosity, surface tension, and other fluid properties. In general,
the droplet generation rate increases with decreasing droplet
diameter, and 1,000,000 droplets per minute is achievable for most
aqueous fluid drops under about 10 .mu.m in diameter.
[0055] Acoustic ejection may be used in any context where precise
placement of a fluid droplet is desirable or necessary. In
particular, the invention may be employed to improve accuracy and
precision associated with nozzleless acoustic ejection. For
example, it is described in U.S. Patent Application Publication No.
20020037579 to Ellson et al. that acoustic ejection technology may
be used to form biomolecular arrays. Similarly, acoustic ejection
technology may be employed to format a plurality of fluids, e.g.,
to transfer fluids from odd-sized bulk containers to wells of a
standardized well plate or to transfer fluids from one well plate
to another. Furthermore, as described in U.S. Patent Application
Publication Nos. 20020109084 and 20020125424, each to Ellson et
al., focused acoustic radiation may serve to eject a droplet of
fluid from a reservoir into any sample vessel for processing and/or
analyzing a sample molecule, e.g., into a sample introduction
interface of a mass spectrometer, an inlet opening that provides
access to the interior region of a capillary, or an inlet port of a
microfluidic device. Similarly, the invention may be used to
transport droplets of analysis-enhancing fluid on a sample surface
in order to prepare the sample for analysis, e.g., for MALDI or
SELDI-type analysis.
[0056] In order to prepare an array on a substrate surface, the
substrate must be placed in droplet-receiving relationship to a
reservoir. Thus, the invention may also employ a positioning means
for positioning the substrate. With respect to the substrate
positioning means and the ejector positioning means, it is
important to keep in mind that there are two basic kinds of motion:
pulse and continuous. For the ejector positioning means, pulse
motion involves the discrete steps of moving an ejector into
position, emitting acoustic energy, and moving the ejector to the
next position; again, using a high performance positioning means
with such a method allows repeatable and controlled acoustic
coupling at each reservoir in less than 0.1 second. A continuous
motion design, on the other hand, moves the ejector and the
reservoirs continuously, although not necessarily at the same
speed, and provides for ejection during movement. Since the pulse
width is very short, this type of process enables over 10 Hz
reservoir transitions, and even over 1000 Hz reservoir transitions.
Similar engineering considerations are applicable to the substrate
positioning means.
[0057] From the above, it is evident that the relative positions
and spatial orientations of the various components may be altered
depending on the particular desired task at hand. In such a case,
the various components of the device may require individual control
or synchronization to direct droplets onto designated sites on a
substrate surface. For example, the ejector positioning means may
be adapted to eject droplets from each reservoir in a predetermined
sequence associated with an array of designated sites on the
substrate surface. Any positioning means of the present invention
may be constructed from, e.g., levers, pulleys, gears, a
combination thereof, or other mechanical means known to one of
ordinary skill in the art.
[0058] FIG. 1 illustrates an exemplary focused acoustic ejection
device suitable for use with the invention, in simplified
cross-sectional view. As with all figures referenced herein, in
which like parts are referenced by like numerals, FIG. 1 is not to
scale, and certain dimensions may be exaggerated for clarity of
presentation. The device 11 includes a plurality of reservoirs,
i.e., at least two reservoirs--a first reservoir indicated at 13
and a second reservoir indicated at 15. Each reservoir contains a
combination of two or more immiscible fluids, and the individual
fluids as well as the fluid combinations in the different
reservoirs may be the same or different. As shown, reservoir 13
contains fluid 14, and reservoir 15 contains fluid 16. Fluids 14
and 16 have fluid surfaces respectively-indicated at 17 and 19. As
shown, the reservoirs are of substantially identical construction
so as to be substantially acoustically indistinguishable, but
identical construction is not a requirement. The reservoirs are
shown as separate removable components but may, if desired, be
fixed within a plate or other substrate. Each of the reservoirs 13
and 15 is axially symmetric as shown, having vertical walls 21 and
23 extending upward from circular reservoir bases 25 and 27 and
terminating at openings 29 and 31, respectively, although other
reservoir shapes may be used. The material and thickness of each
reservoir base should be such that acoustic radiation may be
transmitted therethrough and into the fluid contained within the
reservoirs.
[0059] The device also includes an acoustic ejector 33 comprised of
an acoustic radiation generator 35 for generating acoustic
radiation, and a focusing means 37 for focusing the acoustic
radiation at a focal point near the fluid surface from which a
droplet is to be ejected, wherein the focal point is selected so as
to result in droplet ejection. The focal point may be in the upper
fluid layer or the lower fluid layer, but is preferably just below
the interface therebetween. As shown in FIG. 1, the focusing means
37 may comprise a single solid piece having a concave surface 39
for focusing acoustic radiation, but the focusing means may be
constructed in other ways as discussed below. The acoustic ejector
33 is thus adapted to generate and focus acoustic radiation so as
to eject a droplet of fluid from each of the fluid surfaces 17 and
19 when acoustically coupled to reservoirs 13 and 15, respectively.
The acoustic radiation generator 35 and the focusing means 37 may
function as a single unit controlled by a single controller, or
they may be independently controlled, depending on the desired
performance of the device. Typically, single ejector designs are
preferred over multiple ejector designs, because accuracy of
droplet placement, as well as consistency in droplet size and
velocity, are more easily achieved with a single ejector.
[0060] In operation, each reservoir 13 and 15 of the device is
filled with different fluids, as explained above. The acoustic
ejector 33 is positionable by means of ejector positioning means
43, shown below reservoir 13, in order to achieve acoustic coupling
between the ejector and the reservoir through acoustic coupling
medium 41. If droplet ejection onto a substrate is desired, a
substrate 49 may be positioned above and in proximity to the first
reservoir 13 such that one surface of the substrate, shown in FIG.
1 as underside surface 51, faces the reservoir and is substantially
parallel to the surface 17 of the fluid 14 therein. The substrate
49 is held by substrate positioning means 53, which, as shown, is
grounded. Thus, when the substrate 49 is comprised of a conductive
material, the substrate 49 is grounded as well. Once the ejector,
the reservoir, and the substrate are in proper alignment, the
acoustic radiation generator 35 is activated to produce acoustic
radiation that is directed by the focusing means 37 to a focal
point 55 near the fluid surface 17 of the first reservoir. As a
result, droplet 57 is ejected from the fluid surface 17, optionally
onto a particular site (typically although not necessarily, a
pre-selected, or "predetermined" site) on the underside surface 49
of the substrate. The ejected droplet may be retained on the
substrate surface by solidifying thereon after contact; in such an
embodiment, it is necessary to maintain the substrate surface at a
low temperature, i.e., at a temperature that results in droplet
solidification after contact. Alternatively, or in addition, a
molecular moiety within the droplet attaches to the substrate
surface after contact, through adsorption, physical immobilization,
or covalent binding.
[0061] Then, as shown in FIG. 1B, a substrate positioning means 53
may be used to reposition the substrate 49 (if used) over reservoir
15 in order to receive a droplet therefrom at a second site. FIG.
1B also shows that the ejector 33 has been repositioned by the
ejector positioning means 59 below reservoir 15 and in acoustically
coupled relationship thereto by virtue of acoustic coupling medium
41. Once properly aligned, as shown in FIG. 1B, the acoustic
radiation generator 35 of ejector 33 is activated to produce
acoustic radiation that is then directed by focusing means 37 to a
focal point within the reservoir fluids in reservoir 15, thereby
ejecting droplet 63, optionally onto the substrate.
[0062] It should be evident that such operation is illustrative of
how an acoustic ejector may be used to eject a plurality of
droplets from reservoirs in order to form a pattern, e.g., an
array, on the substrate surface 51. It should be similarly evident
that an acoustic ejector may be adapted to eject a plurality of
droplets from one or more reservoirs onto the same site of the
substrate surface. Furthermore, the ejection of a plurality of
droplets may involve one or more ejectors. In some instances, the
droplets are ejected successively from one or more reservoirs. In
other instances, droplets are ejected simultaneously from different
reservoirs.
[0063] As depicted in FIG. 2, the invention may be used with a
single reservoir to transport fluid into an inlet opening of a
target vessel. Axially symmetric capillary 49 having an inlet
opening 50 disposed on a terminus 51 thereof is provided as a
target vessel. Due to the axial symmetry of the capillary 49, the
inlet opening 50 has a circular cross section. As such, the opening
has a limiting dimension equal to its diameter.
[0064] A hemispherical volume of fluid 14 on a substantially flat
surface 25 of a substrate 13 serves a reservoir. The shape of fluid
14 is a function of the sample wetting properties with respect to
the substrate surface 25. Thus, the shape can be modified with any
of a number of surface modification techniques. In addition, an
ejector 33 is provided comprising an acoustic radiation generator
35 for generating radiation, and a focusing means 37 for directing
the radiation at a focal point near the surface 17 of the fluid 14.
The ejector 33 is shown in acoustic coupling relationship to the
substrate 13 through coupling fluid 41. Proper control of acoustic
wavelength and amplitude results in the ejection of a droplet 57
from the fluid 14 on the substrate 13. As the droplet 57 is shown
having a diameter only slightly smaller than the diameter of the
inlet opening 49, it is evident that this configuration requires
strict control over the droplet size and trajectory.
[0065] The invention is in general a system or method for fluid
transport. A quantity of fluid is held in a reservoir. A droplet
generator is employed to generate droplets from the fluid, for
example a nozzle-based system or a nozzleless system such as an
acoustic ejection system. The droplet generator may or may not make
contact with the fluid in order to generate the droplet. The
droplet generator may be set up to move from one reservoir to
another in order to be able to eject from more than one reservoir.
In a common arrangement, a number of reservoirs form part of an
integral structure, e.g., a well plate, and the structure is moved
by suitable mechanical or electromechanical systems, potentially
under computer control, into a suitable position with respect to
the droplet generator.
[0066] The droplet generator is controlled by a controller,
preferably electronic and connectable to computers through some
communications system. The droplets after generation arrive at a
target, which may be for example a well plate, a porous or
non-porous surface, a substrate, or a structure which is to be
coated by the droplets. The invention also includes a circuit which
controls a characteristic of the generated droplet, increasing the
likelihood that the droplet will not splash or bounce on arriving
at the target. This circuit may be controlled by the controller or
may form part of the controller. The target may already have a
quantity of fluid, and the droplet may coalesce with that quantity
of fluid.
[0067] In general the invention may be used with a range of fluids
and targets. For example, a fluid in the target with which
coalescence could be desired to occur may be of different
composition from the fluid in the droplet. A common compositional
difference encountered in practice is ratio of DMSO to water where
solutions comprising mixtures of DMSO and water are transported.
Compositional differences may lead to differences in physical
parameters such as viscosity and dielectric constant. Thus, for
example, a 70% DMSO/30% water mixture has a viscosity of roughly
3.5 centipoise while an aqueous buffer would have a viscosity of
roughly 1 centipoise. Thus the droplets of the invention may have a
viscosity which is more than 3 times that of the fluid at the
target, or vice versa.
[0068] The fluid with which coalescence could be desired to occur
may be of approximately the same volume as the droplet, for example
another droplet of similar size or two droplets of similar size, or
it may be a larger quantity of fluid, for example a quantity of
fluid more than 100 times larger than the droplet.
[0069] A preferred controller contains a microprocessor with
suitable memory and software or firmware. The microprocessor may be
running an operating system specialized for real time and embedded
applications, as for example QNX from QNX Software Systems (Ottawa,
Ontario, Canada). Some controllers may contain two or more
microprocessors and distribute functions among them as is
convenient from the point of view of the design and operation of
the system. Such a controller is preferably connected to a display
allowing direct operator interaction with the fluid transport
system by pressing of buttons or through a touch screen. Such a
controller is also preferably provided with communications
software, firmware and/or hardware which allows it to communicate
with computers including general-purpose computers which can form
part of an overall laboratory or manufacturing automation network,
and which would allow the controller to operate the fluid transport
system in an automated manner based on commands or information
received from other entities in the overall laboratory or
manufacturing automation system.
[0070] An overall laboratory automation system may include, for
example, a carousel for holding well plates, a robot arm for moving
well plates from one instrument to another, a variety of analytical
instruments and reaction chambers, a pin based fluid transfer
system, and/or an acoustic ejection system. The overall purposes of
the system may include taking quantities of fluids and subjecting
them to analyses (including for example the ascertainment of their
composition and physical properties), reactions designed to produce
particular moieties, and purification steps, all the while
potentially keeping track, by computerized or other means, of the
origin and destination of each fluid in the system and of the
processes and results for each fluid. The system may also be
employed to generate for further use objects which contain or are
coated with fluids moved by the system.
[0071] The tracking of the origin, destination, processes, and
results for each fluid may be performed, for example, by having
controllers such as the fluid transport system controller
communicate that information to a general purpose computer which
stores the information as flat files or in a database. Fluids are
conveniently identified by assigning an identifier to each well
plate in the system and by tracking what is done to each well in
each plate at particular times in a way that allows one to produce
an overall history for the contents of each well of each plate. It
must be kept in mind in this regard that not all changes in fluids
in the system take place as a result of deliberate or planned
action; some may be inevitable changes that occur as a result of
the passage of time, as for example the absorption of water from
the surrounding air or the evaporation of fluids in storage.
[0072] In a laboratory automation system it will generally be
necessary to integrate equipment from different manufacturers. In
this connection the adherence to particular standards may be a
desirable feature of a fluid transport system. Certain fluid
transport systems which form part of a manufacturing environment
may be required to meet further standards relating to manufacturing
as well as being able to support overall system conformance with
the norms of "Good Manufacturing Practice" (GMP) as understood by
the pharmaceutical industry. In particular applications the fluids
being transported may be pathogenic requiring special measures for
their handling which may impact on the design of the
controller.
[0073] A preferred controller of a fluid transport system will also
optionally contain detailed information for achieving efficient
transport of fluid within the system. Such information might be,
for example, preferred ranges of droplet speeds as discussed below,
preferred ranges of Weber numbers, and physical characteristics of
the fluids being manipulated, as for example their conductivity and
permittivity, which are also discussed below, and also their
density, viscosity, surface tension, and the like. Such information
will also include algorithms for operating in a suitably
time-sequenced manner the different actuators of the fluid
transport system, for example well plate transport systems, pin
based fluid transport systems, robot arms, acoustic ejector
transducers, and the electrodes discussed in connection with
certain embodiments below. The time-sequenced algorithms may be
stored in the form of tables which are interpreted by suitable
software or firmware or they may be coded directly as programs or
they may be a mixture of tables and code. The time-sequenced
algorithms may take into account information obtained from various
sensors in the system, for example position and temperature sensors
and digital cameras, as well as stored or measured characteristics
of the fluids being manipulated. Preferably the algorithms may be
modified in the field, e.g., by software downloads over a network,
to take into account the most recent knowledge about the best known
methods for the operation of the fluid transport system. The
controller may also have learning capabilities in which it can,
through analysis of fluids, determine by itself some of the
parameters most suitable for their efficient transport.
[0074] One way to control characteristics of the droplets in a
fluid transport system to reduce the likelihood of splashing or
bounce is to control the speed with which the droplet approaches
the target. The manner in which the speed control is accomplished
may depend on the droplet generation technique.
[0075] Gravity and friction against the surrounding air or other
gas have an impact on droplet speed which has been studied in
connection with the modeling of the behavior of raindrops. Some
control of the impact of these forces is possible, particularly in
the case of downward flow where the drop height can influence the
speed of the droplet at impact. Because it is difficult to zero out
the effect of these forces, their effect may need to be taken into
account in designing systems even where the primary control of
droplet speed is achieved by other means. The system of the
invention encompasses and contemplates the possibility that the
droplet will travel, upon being generated, in a direction generally
against the force of the earth's gravitational attraction, in
particular in a direction 90 to 180 degrees with respect to the
vector which gives the direction of the earth's gravitational
attraction.
[0076] Nanoliter scale droplets of water-like properties have been
studied. For such droplets it is found that speeds which are
preferable for droplet approach with coalescence are in the range
of 0.2 to 10 m/s. A more preferable range is 1.0 to 2.5 m/s, and
within that range a value above 1.5 m/s is particularly
preferred.
[0077] Electric fields may have an impact on the speed of a
droplet, as discussed for example in U.S. Pat. No. 5,541,627 to
Quate, which states generally that water is attracted to an
electric field. This impact may exist even if the droplet is
uncharged if the electric field causes the droplet to become
electrically polarized, as will occur if the droplet consists
primarily of a polarizable solvent such as water.
[0078] It is believed that an uncharged drop will only feel a net
force on it in a nonuniform electric field. In any field, the drop
will develop an induced dipole charge, i.e. a separation of
charges, just as is the case for any dielectric. A dipole in turn
will be attracted toward a concentration of electric field, that is
toward a region of greater field. In a uniform field, the dipole
experiences no net force. Mathematically, the force is proportional
to the gradient of the square of the external electric field.
[0079] When an acoustic ejection system is used for droplet
generation, it is found that the speed of the droplet is
controllable within a range through the amount of acoustic energy
used to cause the ejection to occur. It has been found, for
example, that for 5 nL droplets of 70% DMSO/30% water in an
acoustic ejection system using an F2 lens, control of the speed
between 0.5 and 3 m/s may be achieved readily through control of
acoustic energy above ejection threshold.
[0080] An alternative way to control characteristics of the droplet
to reduce the likelihood of splashing or bounce is to control the
Weber number of the impact. The Weber number is a dimensionless
quantity intended to express the ratio of inertia force to surface
tension force. It is defined as .rho.V.sup.2l/.sigma. where .rho.
is the density of the fluid, V is the velocity, l is a
characteristic length, and .sigma. is surface tension. For a
droplet impact l may be taken to be droplet diameter. Of these
parameters, it may be easier to control V and l. In the literature
there are indications that particular ranges of Weber numbers are
associated with coalescence, at least with the fluids studied in
the particular literature references. Because of this, it may be
desirable for the improvement of coalescence and control of
splashing and bounce to control the parameters that make up the
Weber number of a droplet in order to confine that Weber number
within particular ranges. The particular ranges may vary according
to the composition of the droplets and of the fluid into which the
droplets are intended to merge. The parameters that are controlled
may include the velocity V and the characteristic length l. The
characteristic length l may be controlled, for example, by using
two smaller droplets in place of one larger one, if that would
improve coalescence properties.
[0081] A further preferred way to control characteristics of the
droplet is to generate an electric field in a zone of space through
which the droplet passes. Preferably such an electric field is
generated in the zone of space in which the droplet is
generated.
[0082] An electric field in a zone through which the droplet passes
may be directed with different orientations. It is preferred that
the electric field be oriented along the direction of droplet
separation from a larger mass of fluid.
[0083] An electric field in a zone through which the droplet passes
may be of different magnitudes. For nanoliter scale droplets, it is
preferred that the electric field lie between 1,000 and 100,000
V/m, preferably between 10,000 and 100,000 V/m, more preferably
between 25,000 and 50,000 V/m, and most preferably between 30,000
and 40,000 V/m. To the extent the field is non-uniform, it is
preferred that the field in the zone of space in which the droplet
is generated lie within the preferential ranges just discussed. The
desired intensity of the electric field would preferably be
increased relative to these values if the fluid of the droplet is
only modestly conductive, for example much less conductive than
pure water.
[0084] It is believed that an electric field component in the
direction of droplet separation from a larger mass of fluid may
cause the droplet to have an electric charge when it is separated,
and that the existence of such electric charges can, if they are of
appropriate magnitudes, facilitate droplet coalescence. The
mechanism of facilitation is not clearly understood, but it is
believed to be an effect above and beyond the possible acceleration
of the droplet by the electric field. A mechanism which has been
hypothesized is an effective change in the droplet's surface
tension with the free charge present. The surface tension is
reduced because the charges on the drop surface repel one
another.
[0085] With certain forms of droplet generation (e.g., acoustic
ejection), droplet separation from a mass of fluid is commonly
preceded by the formation of a projection from the fluid surface.
For a fluid of some degree of conductivity, in the presence of an
electric field, such a projection should become charged so as to
maintain it at an approximate electric equipotential with the mass
of fluid. It is hypothesized that as the droplet forms from the
projection, the projection is essentially pinched off, and the net
charge developed along the surface of the projection is carried
away with the drop. The charge on the droplet may be approximately
described by the relation Q.about.4.pi..epsilon..sub.0ahE where a
denotes the droplet radius, h denotes the height of the mound at
the time of droplet break-off, and E denotes the electric field
intensity. In acoustic ejection, for DMSO/water mixtures the height
h has been observed to be approximately 5 times the drop diameter,
so that h.about.10a. An empirical study of the charge on
acoustically ejected droplets is presented as part of Example 1
below.
[0086] It is believed that an electric field component in the
direction of droplet arrival at the target may assist in the
coalescence on account of the polarization which the droplet will
experience. In general, a dielectric in the presence of an external
electric field will develop a dipole moment, corresponding to a
separation of charge along the direction of the external field.
This redistribution of charges counteracts the external field in
such a way as to cause the net electric field strength inside the
dielectric to be smaller than the external field strength, and in
the limiting case of a perfectly conductive material, to be zero.
In the particular case of a droplet approaching and potentially
coalescing with a mass of fluid, if there is an electric field in
the direction of approach, the induced dipole moment of the droplet
will cause it to be attracted to the fluid mass.
[0087] Two limiting cases of this occur if the fluid mass is either
(1) of the same size as the approaching droplet, or (2) is much
larger than the approaching droplet, so that it can be considered
an infinite half-space. These cases are very similar, especially
for a relatively conducting fluid, which in practice is common. For
the case in which the mass of fluid is very large, the induced
dipole moment of the ejected droplet can be considered to generate
a mirror dipole moment within the fluid mass, consistent with the
method of images well known in electrostatics, as depicted in FIG.
3. Treating the polarization of the droplet as that of a spherical
conductor (a reasonable approximation for high-dielectric-constant
fluids), the dipole moment of the ejected drop may be written as:
p=4.pi..epsilon..sub.0Ea.sup.3 where .epsilon..sub.0 denotes the
permittivity of free space, E denotes the applied external electric
field in the vicinity of the target, and a is the radius of the
drop. For simplicity, we consider the geometry of FIG. 3, where z
is in the (downward) vertical direction, and the target is taken to
be at z=0. The z-component of the electric field due to the "image"
(upper) droplet, at the position of the arriving droplet, a height
z below the fluid surface, can be written as:
E.sub.z=1/4Ea.sup.3/z.sup.3 It follows that the force on the
arriving droplet, due to the interaction of its dipole moment with
this electric field is: F.sub.z=pdE.sub.z/dz=
3/2.pi..epsilon..sub.0E.sup.2a.sup.6/z.sup.4 This force is directed
upward, attracting the arriving droplet toward the surface of the
fluid at the target. The force, which varies as the inverse fourth
power of the height of the droplet below the fluid surface, will be
strongest when the droplet is very near the surface of the
receiving fluid. When the droplet just "touches" the receiving well
fluid, i.e. when z=a, the dipole force is F.sub.z=
3/2.pi..epsilon..sub.0E.sup.2a.sup.2
[0088] With an electric field of 30,000 V/m and 2.5 nl drops, the
associated drop radius is 84 .mu.m and the dipole force with z=a is
approximately F.sub.z.about.3.times.10.sup.-10 N. This may be
compared to the gravitational force on the drop mg
.about.2.6.times.10.sup.-8 N. Thus, the dipole force is only about
one hundredth that of gravity.
[0089] For the other limiting case, in which the receiving mass of
fluid is of the same size as the ejected droplet, the analysis is
very similar to that above. In this case, the receiving fluid mass
forms a physical dipole moment equivalent to that of the drop, in
the presence of the external electric field. Thus, what was
previously an image dipole at a distance 2z from the ejected drop,
is now an equivalent dipole, at a distance z from the drop. Hence
the dipole-dipole force between the ejected drop and the receiving
fluid mass is 8 times larger than that for the previous case, when
the ejected drop is a distance z from the fluid mass. On the other
hand, the limiting distance between the ejected drop and receiving
fluid mass is now 2a. Thus the maximum force between the ejected
drop and receiving fluid mass is half that described previously for
the semi-infinite fluid mass. It is seen therefore that the
polarization effects responsible for attraction of ejected drop to
the receiving fluid mass are of the same order of magnitude, for
the two limiting cases.
[0090] When using an electric field to reduce splash and bounce
and/or improve droplet coalescence, it may be desirable that the
electric field not substantially affect the path followed by
droplet prior to its arrival at the target. It may even be
desirable that the electric field not substantially alter the speed
of the droplet, effecting for example a change of no more than 10%
in the velocity. Such relative independence of the path and speed
from electric field may be helpful, for example, in order to make
it easier to ensure that successive droplets aimed at the same
place on the target arrive at that place on the target repeatably.
If the fluid on the target is itself being deposited droplet by
droplet, as where the fluid transfer system is transferring a
larger quantity of fluid as a series of droplets, such
repeatability may be desirable to facilitate coalescence. Studies
in the literature such as that authored by Professor Orme cited
above indicate that the impact parameter has been seen as being of
importance in coalescence of raindrops. The independence of path
and speed from electric field may also be helpful simply in order
to make other parameters of the total fluid transport system
settable independently of the electric field.
[0091] A common way to generate an electric field in a region of
space is to have two electrodes A and B which are held at different
potentials, such that the region of space overlaps with the region
between the electrodes. The electrodes may have a wide variety of
shapes, for example, flat sheets of solid sufficiently conducting
material, material of low conductivity coated or laminated with
material of higher conductivity, rows of wires, and grids of wires.
There is considerable freedom in the placement of the electrodes,
for example to avoid obstructing the flow of fluid or the movement
of reservoirs and targets. Alternatively, as in examples 3 and 4
below, an electrode may be placed in the path of the fluid with
holes to allow the fluid to pass through.
[0092] A voltage may be permanently wired into the system and the
electrodes energized at the fixed voltage whenever the system is
powered on. This would make sense, for example, if a particular
electrode voltage is found to be advantageous during generation of
droplets in the operating range in which a fluid transfer system of
the invention is expected to be used. Alternatively, and for
greater flexibility, it is possible to make the voltages settable,
for example, through the controller of the fluid transport system,
or through one or more external inputs to the circuit producing the
electric field. Under certain circumstances it can be advantageous
to have a time varying voltage, for example a voltage which changes
during different stages of the generation of each droplet, or a
voltage which is held at a constant value for a predetermined time,
for example during droplet generation, and zero otherwise.
[0093] As is discussed in detail in U.S. patent application Ser.
No. 10/340,557 to Mutz et al., various phenomena exist which can
cause droplets to have uncontrolled charge. It is possible to deal
with uncontrolled charge in various ways which involve in general
terms providing a conducting path, or providing ionization, so that
the fluid from which droplets are formed discharges. For example,
the reservoirs in which the fluid is held might be made from a
sufficiently conducting material and grounded.
[0094] In connection with the embodiment of this invention in which
the droplet characteristics are controlled by means of an electric
field, it is found to be helpful to also use measures for the
reduction of electric charge of the general type described in
application Ser. No. 10/340,557. It is particularly helpful to have
the reservoir holding fluid from which droplets are made be an
approximate or exact ground, using techniques discussed in that
application such as ionization discharging. In that case, a single
electrode held at a nonzero voltage with respect to ground can
create the electric field which is used for droplet control.
[0095] The voltages of the electrodes which are used to create the
electric field can be adjusted by those of skill in the art to
achieve a desired field intensity. The adjustment can be, for
example, purely experimental. It can alternatively be based on well
known formulas (e.g., for the electric field between parallel
plates) as found for example in books on electromagnetics such as
J. D. Jackson, Classical Electrodynamics (2d ed. 1975). It can
alternatively be based on well known numerical techniques for
computing electrostatic fields by numerical solution of Laplace's
equation.
[0096] Alternatively, the voltages of the electrodes may be
adjusted in order to optimize an experimental measure of splash,
bounce, or coalescence. For example, one could run a number of
ejections against suitable targets and use a digital camera to
capture images of the process of droplet impact on the fluid at the
targets for human or automatic analysis. Alternatively, one could
use methods for scoring droplet coalescence from visual examination
of the target after impact, for example by looking for multiple
droplets where there should only be one if coalescence
occurred.
[0097] FIG. 4 depicts a particular embodiment of the circuit of the
invention which is preferred. The fluid transport system in which
this embodiment is used is an acoustic ejection system with
droplets ejected upwards from a source reservoir to a target. In
this embodiment the electrode is in the form of a grid, which is
located behind the target. The source reservoir is maintained as an
approximate ground by means of ionization, by the existence of true
grounds in the vicinity, and/or by electrical conduction through
the acoustic coupling medium.
[0098] As may be seen, the grid is driven by an N-type MOSFET 12
through a 5 M.OMEGA. resistor 14. When the voltage on the gate of
the N-type MOSFET is high (above threshold), the grid is connected
to ground via the 5 M.OMEGA. resistor 14. When the gate voltage is
driven low, the grid is connected via resistor 14 and another 5
M.OMEGA. resistor 16 to the output of a DC to DC converter 18 which
converts 0 to 5 V magnitudes to 0 to 1500 V. The voltage input of
the DC to DC converter 18 is in turn connected to a variable
voltage source 20 producing 0 to 5 V.
[0099] The gate of the N-type MOSFET 12 is connected via a 10
k.OMEGA. resistor 22 to the output of the variable voltage source
20. The gate is also driven by the output of a NAND gate 24 which
allows two separate external control inputs 26 and 28 to set the
gate low and thus disconnect the grid from ground, connecting it
instead to the output of the DC to DC converted 18.
[0100] External logic (not shown) could employ the external inputs
26 and 28 to ensure that when the fluid transport system is loading
well plates or the door is open, the grid is switched to ground via
the N-type MOSFET 12. When droplet ejection starts, the control
inputs could be used to bring the grid up to a fixed voltage
between 500 and 1500 volts DC, with 800 V preferred. This voltage
is applied evenly to the entire grid. This creates a weak static
field that extends from the target plate to approximate ground
planes in the drop ejection chamber and in the coupling fluid under
the source plate wells. After drop transfer, the grid could be then
switched back to ground. The grid is either completely on, or
completely off.
[0101] As will be understood by those of skill in the art, the
components in FIG. 4 would also have power supply connections
which, as is common in electronics, are not explicitly shown.
[0102] In a variant on this preferred embodiment, the voltage
alternates between positive and negative levels in order to prevent
charge buildup on the target wells. Variants are also possible
where the voltage is turned on during well plate loading and where
the grid is left floating for some portion of the ejection
cycle.
[0103] In an alternative preferred embodiment, an ionizer which
creates a biased ion cloud is used to charge target well plates as
they enter the system. The biased ion cloud is created, for
example, by driving the ionizer with a suitably asymmetric or
biased waveform. This ionizer could make use of standard ionization
bars such as those made by Julie Industries (Wilmington, Mass.).
The target well plate is then electrically isolated during the
ejection cycle. The electric field modifying coalescence properties
would be caused chiefly by the charge which was imparted to the
target well plate by means of the biased ion cloud ionizer. Instead
of imparting charge to the target well plate, it would be possible
to charge an object placed near the target well plate as an
alternative way of generating an electric field.
[0104] In another alternative preferred embodiment, a corona
discharge which produces ions of a particular polarity in a
localized portion of space may be used to charge a droplet.
Preferably a stream of gas directs the ions produced by the corona
discharge to the droplet. Alternatively, the droplet may pass
through the discharge. In this case, preferably the corona
discharge is stable, as for example between a pin and a plane.
[0105] Where an electric field is employed to improve droplet
coalescence and that electric field charges the droplets, the
transfer of a large number of such droplets into a single mass of
fluid at the target may produce a significant net charge in that
mass of fluid, which would tend to screen an applied electric field
and reduce its beneficial effects. One can estimate the field that
might be produced by the deposition of the droplet charge in the
receiving mass of fluid. That charge would tend to spread across
the receiving fluid meniscus as a surface charge sheet, so that if
the diameter of the fluid is d, the charge density of the sheet
would be .sigma.=NQ/d.sup.2 where N is the number of droplets
transferred and Q is the charge per individual droplet. If we
assume that the direction of droplet flight between source
reservoir and target is the vertical z direction, that the field is
produced by an electrode at voltage Vabove the target, and that the
source is an approximate ground, we get the equation
E=.sigma./.epsilon..sub.0+[V/z.sub.tf)]/[1+(.epsilon..sub.0/.epsilon..sub-
.tf)(z.sub.elect/z.sub.tf-1)] Here z.sub.elect denotes the height
of the electrode above the source fluid, z.sub.tf denotes the
height of the target fluid meniscus above the source fluid, and
.epsilon..sub.tf represents the permittivity of the target fluid.
For any aqueous target fluid, the quantity
.epsilon..sub.0/.epsilon..sub.tf would be much less than unity, and
typically z.sub.elect/z.sub.tf would be somewhat greater than
unity. Thus, the quantity
(.epsilon..sub.0/.epsilon..sub.tf)(z.sub.elect/z.sub.tf-1)<<1,
so that
1/[1+(.epsilon..sub.0/.epsilon..sub.tf)(z.sub.elect/z.sub.tf-1)].abo-
ut.1-(.epsilon..sub.0/.epsilon..sub.tf)(z.sub.elect/z.sub.tf-1),
and the electric field may be approximated as:
E.about.(V/z.sub.tf)[1-(.epsilon..sub.0/.epsilon..sub.tf)(z.sub.elect/z.s-
ub.tf-1)]+(.sigma./.epsilon..sub.tf)(z.sub.elect/z.sub.tf-1)
[0106] The first term of this expression,
E.sub.init=(V/z.sub.tf)[1-(.epsilon..sub.0/.epsilon..sub.tf)(z.sub.elect/-
z.sub.tf-1)], is the field between source and target before any
droplets arrive. The second term of this expression
E.sub.drop=(.sigma./.epsilon..sub.tf)(z.sub.elect/z.sub.tf-1) is
the portion of the electric field due to the deposited charge at
the meniscus. It is preferable, to avoid uncontrolled change in the
actual electric field E, for E.sub.drop to be considerably less
than E.sub.init, for example ten times less. Applying again the
fact that
(.epsilon..sub.0/.epsilon..sub.tf)(z.sub.elect/z.sub.tf-1)<<1,
we have
E.sub.drop/E.sub.init.about.(.sigma./.epsilon..sub.tf)(z.sub.elect--
z.sub.tf)/V
[0107] If Q=1.times.10.sup.-13 C, and d=3.5 mm, we have
.sigma.=8.16.times.10.sup.-9 C/m.sup.2 per droplet. If
z.sub.elect-z.sub.tf=5 mm, .epsilon..sub.tf=80.epsilon..sub.0, and
V=800 V, then the ratio
E.sub.drop/E.sub.init.about.(.sigma./.epsilon..sub.tf)(z.sub.elect-z.sub.-
tf)/V=76.times.10.sup.-6 per droplet. Thus, to keep this ratio
below 1/10, it is desirable to deposit no more than 1300 droplets.
If it is desired to deposit more than that number of droplets, it
might be worthwhile reversing the direction of the electric field
by reversing the polarity of V, so that charge of opposite polarity
starts to neutralize the accumulated .sigma. on the surface of the
target fluid.
[0108] Variations of the present invention will be apparent to
those of ordinary skill in the art. For example, the invention may
be suitable for use with any of the performance enhancing features
associated with acoustic technologies such those described in U.S.
patent application Ser. Nos. 10/010,972, and 10/310,638, each
entitled "Acoustic Assessment of Fluids in a Plurality of
Reservoirs," filed Dec. 4, 2001 and Dec. 4, 2002, respectively,
inventors Mutz and Ellson and U.S. patent application Ser. No.
10/175,375, entitled "Acoustic Control of the Composition and/or
Volume of Fluid in a Reservoir," filed Jun. 18, 2002, inventors
Ellson and Mutz. In addition, the invention may be used in a number
of contexts such as handling pathogenic fluids (see U.S. patent
application Ser. No. 10/199,907, entitled "Acoustic Radiation of
Ejecting and Monitoring Pathogenic Fluids," filed Jul. 18, 2002,
inventors Mutz and Ellson) and manipulating cells and particles
(see U.S. Patent Application Publication Nos. 20020090720 and
20020094582).
[0109] It is to be understood that while the invention has been
described in conjunction with the preferred specific embodiments
thereof, that the foregoing description and the examples that
follow are intended to illustrate and not limit the scope of the
invention. Other aspects, advantages, and modifications within the
scope of the invention will be apparent to those skilled in the art
to which the invention pertains.
[0110] All patents, patent applications, and publications mentioned
herein are hereby incorporated by reference in their entireties.
However, where a patent, patent application, or publication
containing express definitions is incorporated by reference, those
express definitions should be understood to apply to the
incorporated patent, patent application, or publication in which
they are found, and not to the remainder of the text of this
application, in particular the claims of this application.
[0111] The following examples are put forth so as to provide those
of ordinary skill in the art with a complete disclosure and
description of how to implement the invention, and are not intended
to limit the scope of what the inventors regard as their
invention.
EXAMPLE 1
[0112] Within an acoustic ejection system, a conducting plate was
situated 7 mm above a 384 well plate in the source position (wells
facing up). The 384 well plate contained 30 .mu.L of 70% DMSO/30%
water in all wells. The ejection of 2.5 nL drops was then captured
on video, with 0 V and with 800 V applied to the upper plate. The
acoustic coupling fluid, which contacts the bottom of the well
plate, was held in contact with ground potential. The electric
field with the electrode at 800V is estimated to be 49,000 V/m
based on a distance of 16.5 mm between the ground plane and the
electrode. With electric field, the vertical position of the 2.5 nL
drops was .DELTA.z=61 .mu.m higher at t=6.6 ms after ejection,
compared to having the upper electrode grounded.
[0113] It is hypothesized that the difference of position caused by
the electric field was due to the droplet becoming charged during
the ejection. To calculate the charge q imparted to the droplet we
proceed as follows.
[0114] The forces on a droplet of charge q at position z and time t
are -gm for gravity, qE for electric field, and, if Stokes' law
applies, -kv(t) for drag where k=3.pi..eta.d, .eta. being the
viscosity of air, 1.8.times.10.sup.-5 kg-s/m, and d being the
droplet diameter. Applying Newton's law, we get
md.sup.2z/dt.sup.2=-gm+qE-k(dz/dt) which can be rewritten as
mdv/dt=-gm+qE-kv where v=dz/dt is the velocity of the droplet in
the z direction.
[0115] In general, a linear first order differential equation
dv/dt+Av+B=0 with A.noteq.0 has the solution
v(t)=(v.sub.0+B/A)exp(-tA)-B/A where v(0)=v.sub.0. Integrating,
z(t)=(1/A)(v.sub.0+B/A)(1-exp(-tA))-Bt/A if z=0 at I=0. Here A=k/m
and B=g-qE/m. The difference between z(t) with the electric field E
on and off is thus .DELTA. .times. .times. z = ( 1 / A 2 ) .times.
( qE / m ) .times. ( 1 - exp .function. ( - tA ) ) - ( qE / m
.times. .times. A ) .times. t = ( 1 / A 2 ) .times. ( qE / m )
.times. ( 1 - exp .function. ( - tA ) - tA ) , ##EQU1## so that
q=.DELTA.z(m/E)(A.sup.2/(1-exp(-tA)-tA)). When A.fwdarw.0 (meaning
the drag force .about.0), taking limits in the formula for q, we
get the simpler formula q=2.DELTA.z(m/Et.sup.2).
[0116] The value m is about 2.7 .mu.m given that the drop has a 2.5
nL volume and 70% DMSO/30% water has a density of about 1.07.
Values for E have been given above. The droplet diameter is
.about.168 .mu.m for a 2.5 nL sphere. We thus have A.about.10
s.sup.-1 so that tA.about.0.066 (dimensionless). Substituting these
values in the formula for q given above we get
q.about.1.6.times.10.sup.-13 C.
[0117] The difference in velocities with and without the electric
field is given by .DELTA.v=qE/A(1-exp(-tA)).about.0.02 m/s. As may
be seen, in this example the electric field had little effect on
the velocity of the droplet, which is on the order of 2 m/s.
EXAMPLE 2
[0118] A study was carried out with an acoustic ejection system to
better understand the influence of droplet speed on coalescence. A
Krautkramer 15 MHz F2 lens was used, and 70% DMSO/30% water was
ejected into an inverted target well, filled with NaOH buffer. The
receiving fluid surface was slightly convex. Drops were ejected of
nominal volume 5 nL, 2.5 nL, and 1.25 nL.
[0119] First, the drop volumes and velocities were extracted from
video data, as a function of power above threshold. These are shown
in FIGS. 5A through 5C.
[0120] In these graphs, the displayed power is based on the
amplitude delivered to the acoustic transducer, which may be
subject to saturation in the ranges indicated. The data span the
entire range from ejection threshold (0 dB always corresponds to
threshold) to satellite threshold, so that the data should
represent well the operating window for each ejection condition.
The drop velocities quoted in FIGS. 5A-5C are the initial
velocities, directly above the source fluid.
[0121] FIGS. 6A-6C show curves representing the drop bounce
probability (from the filled 384 well), as a function of the
incident drop velocity at the destination well. The velocities in
these figures are velocities upon arrival at the destination
well.
EXAMPLE 3
[0122] In a fluid transfer system based on acoustic ejection
between well plates and having an electrode behind the target
plate, metal foil was placed over the source plate, and holes were
punctured to access the wells. This metal aperture over the source
plate was then grounded, and acted to screen the applied electric
field from the source well fluid (the field strength with the foil
present would be less than 1% of that present without the foil).
Thus there would be minimal droplet charging with the foil in
place. With the foil present, the field between the source plate
and the destination well fluid would actually be increased,
however, so that a dipole-dipole interaction would be enhanced. It
was found that with the foil present, there was essentially no
reduction in droplet bounce when the 800V was applied to the grid
behind the target plate.
EXAMPLE 4
[0123] A foil with apertures was used in the system of Example 3,
but this time over the target plate rather than over the source
plate. The foil was held at 800 V, as was the grid behind the
target plate. With this arrangement, there is essentially no
electric field present within the target plate wells, but there is
drop charging, due to the electric field inside the source plate.
It was found that for this scenario, there was excellent reduction
in drop bounce.
* * * * *