WO2014078100A1 - Mécanismes et procédés de chargement d'un actionneur de gouttelettes avec un fluide de remplissage - Google Patents

Mécanismes et procédés de chargement d'un actionneur de gouttelettes avec un fluide de remplissage Download PDF

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Publication number
WO2014078100A1
WO2014078100A1 PCT/US2013/068080 US2013068080W WO2014078100A1 WO 2014078100 A1 WO2014078100 A1 WO 2014078100A1 US 2013068080 W US2013068080 W US 2013068080W WO 2014078100 A1 WO2014078100 A1 WO 2014078100A1
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WO
WIPO (PCT)
Prior art keywords
fluid
droplet
actuator
reservoir
droplet actuator
Prior art date
Application number
PCT/US2013/068080
Other languages
English (en)
Inventor
Donovan E. BORT
Ramakrishna Sista
Yan-You Lin
Christopher D. SANSEVIERI
Jarred DE VILLE
Original Assignee
Advanced Liquid Logic, Inc.
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Advanced Liquid Logic, Inc. filed Critical Advanced Liquid Logic, Inc.
Publication of WO2014078100A1 publication Critical patent/WO2014078100A1/fr

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Classifications

    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L3/00—Containers or dishes for laboratory use, e.g. laboratory glassware; Droppers
    • B01L3/50—Containers for the purpose of retaining a material to be analysed, e.g. test tubes
    • B01L3/502—Containers for the purpose of retaining a material to be analysed, e.g. test tubes with fluid transport, e.g. in multi-compartment structures
    • B01L3/5027—Containers for the purpose of retaining a material to be analysed, e.g. test tubes with fluid transport, e.g. in multi-compartment structures by integrated microfluidic structures, i.e. dimensions of channels and chambers are such that surface tension forces are important, e.g. lab-on-a-chip
    • B01L3/502769—Containers for the purpose of retaining a material to be analysed, e.g. test tubes with fluid transport, e.g. in multi-compartment structures by integrated microfluidic structures, i.e. dimensions of channels and chambers are such that surface tension forces are important, e.g. lab-on-a-chip characterised by multiphase flow arrangements
    • B01L3/502784—Containers for the purpose of retaining a material to be analysed, e.g. test tubes with fluid transport, e.g. in multi-compartment structures by integrated microfluidic structures, i.e. dimensions of channels and chambers are such that surface tension forces are important, e.g. lab-on-a-chip characterised by multiphase flow arrangements specially adapted for droplet or plug flow, e.g. digital microfluidics
    • B01L3/502792—Containers for the purpose of retaining a material to be analysed, e.g. test tubes with fluid transport, e.g. in multi-compartment structures by integrated microfluidic structures, i.e. dimensions of channels and chambers are such that surface tension forces are important, e.g. lab-on-a-chip characterised by multiphase flow arrangements specially adapted for droplet or plug flow, e.g. digital microfluidics for moving individual droplets on a plate, e.g. by locally altering surface tension
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2200/00—Solutions for specific problems relating to chemical or physical laboratory apparatus
    • B01L2200/02—Adapting objects or devices to another
    • B01L2200/026—Fluid interfacing between devices or objects, e.g. connectors, inlet details
    • B01L2200/027—Fluid interfacing between devices or objects, e.g. connectors, inlet details for microfluidic devices
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2200/00—Solutions for specific problems relating to chemical or physical laboratory apparatus
    • B01L2200/06—Fluid handling related problems
    • B01L2200/0642—Filling fluids into wells by specific techniques
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2200/00—Solutions for specific problems relating to chemical or physical laboratory apparatus
    • B01L2200/06—Fluid handling related problems
    • B01L2200/0684—Venting, avoiding backpressure, avoid gas bubbles
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2400/00—Moving or stopping fluids
    • B01L2400/04—Moving fluids with specific forces or mechanical means
    • B01L2400/0403—Moving fluids with specific forces or mechanical means specific forces
    • B01L2400/0415—Moving fluids with specific forces or mechanical means specific forces electrical forces, e.g. electrokinetic
    • B01L2400/0427—Electrowetting
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L3/00—Containers or dishes for laboratory use, e.g. laboratory glassware; Droppers
    • B01L3/02—Burettes; Pipettes
    • B01L3/0275—Interchangeable or disposable dispensing tips

Definitions

  • the invention relates to mechanisms and methods of loading a droplet actuator with filler fluid to reduce or eliminate the trapping of air in a droplet operations gap of the droplet actuator during the loading process.
  • a droplet actuator typically includes one or more substrates configured to form a surface or gap for conducting droplet operations.
  • the one or more substrates establish a droplet operations surface or gap for conducting droplet operations and may also include electrodes arranged to conduct the droplet operations.
  • the droplet operations substrate or the gap between the substrates may be coated or filled with a filler fluid that is immiscible with the liquid that forms the droplets.
  • the invention is directed to mechanisms and methods of loading a droplet actuator with filler fluid to reduce or eliminate the trapping of air in a droplet operations gap of the droplet actuator during the loading process.
  • the invention provides a mechanism that includes: a droplet actuator, in which the droplet actuator includes a bottom substrate and a top substrate, in which the bottom substrate and the top substrate are separated by a droplet operations gap, in which the top substrate includes a filler fluid loading port, further in which the filler fluid loading port provides a fluid path through an opening in the top substrate to the droplet operations gap; and a filling fixture configured for controlled loading of the droplet actuator with filler fluid; in which the controlled loading of the droplet actuator with filler fluid includes loading filler fluid in the droplet actuator at a speed that is sufficiently slow to reduce or eliminate the trapping of air in the droplet operations gap of the droplet actuator.
  • the controlled loading of the droplet actuator with filler fluid further may include loading the filler fluid into the droplet operations gap of the droplet actuator at a speed that is sufficiently slow to reduce or eliminate loss of filler fluid through one or more openings in the mechanism.
  • the controlled loading of the droplet actuator with filler fluid further may also include loading the filler fluid into the droplet operations gap of the droplet actuator at a speed that is rapid enough for practical use.
  • the invention may include a filler fluid loading port that includes a cylindrical element around the opening in the top substrate, in which the cylindrical element protrudes outwardly from the top substrate and away from the droplet operations gap.
  • the cylindrical element may include an alignment feature configured to allow the filling fixture to be affixed thereto.
  • the filling fixture of the invention may be mechanically and fluidly coupled to the filler fluid loading port.
  • the filling fixture may further include a filling fixture fluid reservoir, such as a cylinder-shaped or dish-shaped filling fixture fluid reservoir.
  • the filling fixture fluid reservoir may also include a floor (i.e., a bottom surface), in which the floor includes an outlet, and further in which the outlet includes an opening in the floor.
  • An outlet shroud may also be positioned around the outlet, e.g., such that the outlet shroud is mechanically coupled to the filler fluid loading port of the top substrate of the droplet actuator.
  • the outlet shroud may also be configured to be press-fitted onto the filler fluid loading port of the top substrate of the droplet actuator.
  • the outlet may be configured to communicate filler fluid from the filling fixture to the droplet operations gap of the droplet actuator.
  • the outlet of the filling fixture may also be aligned with the filler fluid loading port in the top substrate of the droplet actuator.
  • the outlet of the filling fixture may also be fluidly coupled to the filler fluid loading port in the top substrate of the droplet actuator.
  • the filling fixture, the opening in the floor of the filling fixture fluid reservoir, the outlet, and the outlet shroud or the outlet tip may be substantially centered with respect to the filling fixture fluid reservoir.
  • the outlet and the outlet shroud or the outlet tip may also be positioned substantially offset from the center of the filling fixture fluid reservoir.
  • a majority of the filling fixture fluid reservoir may be positioned off the edge of the droplet actuator when in use.
  • the filling fixture may also be configured to allow visual observation of the loading the filler fluid into the droplet operations gap of the droplet actuator.
  • the filling fixture fluid reservoir may also be kidney-shaped or piano-shaped.
  • the invention may include an outlet tip configured to be fitted into the filler fluid loading port of the top substrate of the droplet actuator.
  • the outlet tip may include a tapered pipette-like tip.
  • the filling fixture fluid reservoir may be trough-shaped.
  • the outlet tip may include a two-piece pipette.
  • the two-piece pipette may include a first pipette tip fitted into a second pipette tip.
  • the first pipette tip and the second pipette tip each may include a hollow tapered tube that is open at each end, the first pipette tip and the second pipette tip each further including an inlet and an outlet, in which the inlet has a larger diameter than the outlet.
  • the first pipette tip and the second pipette tip are each formed of molded plastic.
  • the first pipette tip may be configured to be press fitted or snap fitted into the inlet of the second pipette tip.
  • the first pipette tip may include a 1-ml-pipette tip, and in which the second pipette tip may include a 30 ⁇ 1-pipette tip, in which the inner diameter of the outlet of the second pipette tip may be about 0.02 inches.
  • the first pipette tip and the second pipette tip are configured to allow the filler fluid to fill the droplet operations gap of the droplet actuator in a manner comparable to a medical drip tube.
  • the two-piece pipette may include a transparent, semi-transparent, or opaque material.
  • the filling fixture and the two-piece pipette are formed by machining, injection molding, or casting.
  • materials suitable for forming the filling fixture and the two-piece pipette by injection molding comprise a material selected from the group consisting of polyoxymeihylene, polymethyl methacrylate (PMMA), polycarbonate (PC), polypropylene (PP), and polystyrene (PS).
  • the opening in the top substrate of the invention may include a diameter dl .
  • the filler fluid loading port of the invention may also include an inside diameter, in which the inside diameter may include the diameter dl .
  • the filler fluid loading port may include an outside diameter, in which the outside diameter may include a diameter d2.
  • the diameter d2 may be from about .09 inches to about .13 inches, or the diameter d2 may be about .1 1 inches.
  • the diameter dl may be from about .01 13 inches to about .0913 inches, or the diameter dl may be from about .0413 inches to about .0613 inches, or the diameter dl may be about .0513 inches.
  • the opening in the floor of the filling fixture fluid reservoir may include a diameter d3.
  • the diameter d3 may be from about 0.001 inches to about 0.1 inches, or the diameter d3 may be from about 0.01 inches to about 0.06 inches, or the diameter d3 may be from about 0.02 inches to about 0.05 inches, even or the diameter d3 may be from about .04 inches to about .05 inches, and yet or the diameter d3 may be about 0.047 inches.
  • the filling fixture fluid reservoir of the invention may be configured to hold a volume VI of filler fluid.
  • the volume VI may be from about 1 ml to about 1.5 ml, or the volume VI may be about 1.26 ml.
  • the filling fixture fluid reservoir may also be configured to comprise a height h.
  • the height h may be from about 7.5 mm to about 8.5 mm, or the height h may be about 8 mm.
  • the floor of the filling fixture fluid reservoir may be substantially flat, or the floor of the filling fixture fluid reservoir may be counter-sunk leading to the opening in the floor of the filling fixture fluid reservoir.
  • the outlet shroud of the invention may include an inside diameter, in which the inside diameter of the outlet shroud may be at least slightly greater than the outside diameter d2 of the filler fluid loading port.
  • the inside diameter of the outlet shroud may also be configured to allow the outlet shroud to be press fitted onto the filler fluid loading port.
  • the filling fixture of the invention may be configured for loading the filler fluid into the droplet operations gap of the droplet actuator by gravity feed.
  • the filling fixture may also be configured to provide a funnel-like function.
  • the filler fluid may include a low- viscosity oil, e.g., in which the low viscosity oil may be selected from the group consisting of silicone oil and hexadecane oil.
  • the droplet actuator of the invention may include a plurality of on- actuator reservoirs.
  • One or more of the on-actuator reservoirs may be configured to process a sample fluid and/or a reagent fluid.
  • the top substrate of the droplet actuator may include a plurality of loading ports, further in which each of the plurality of loading ports provides a fluid path through the top substrate to one or more of the on-actuator reservoirs.
  • Each of the plurality of on-actuator reservoirs may be in fluid communication with the droplet operations gap of the droplet actuator.
  • the droplet actuator may include electrical contact points, and the filling fixture may be physically connected to the droplet actuator either at a corner of the droplet actuator opposite the electrical contact points or at a corner of the droplet actuator including the electrical contact points.
  • the invention provides a method for controlled loading of a droplet actuator with filler fluid, the method including use of any of the mechanisms disclosed herein to load the filler fluid in the droplet actuator at a speed that may be sufficiently slow to reduce or eliminate the trapping of air in the droplet operations gap of the droplet actuator.
  • the method may include use of any of the mechanisms disclosed herein to load the filler fluid in the droplet actuator at a speed that may be sufficiently slow to reduce or eliminate loss of filler fluid through one or more openings in the mechanism.
  • the method may further include use of any of the mechanisms disclosed herein to load the filler fluid in the droplet actuator at a speed that may be rapid enough for practical use.
  • the invention provides a method for controlled dispensing of fluid into a droplet operations gap of a droplet actuator including a plurality of on-actuator reservoirs, the method including: electrowetting-mediated dispensing of fluid from a first on-actuator reservoir, including dispensing the fluid from the first on-actuator reservoir using a first set of reservoir electrodes; and electrowetting-mediated retention of fluid from one or more additional on- actuator reservoirs, including retaining the fluid from the one or more additional on-actuator reservoirs by using one or more additional sets of reservoir electrodes; in which the controlled dispensing of fluid reduces or eliminates flooding the droplet operations gap of the droplet actuator with fluid from the one or more additional on-actuator reservoirs.
  • the plurality of on- actuator reservoirs may include an array of on-actuator reservoirs.
  • the one or more additional on-actuator reservoirs may be adjacent to the first on-actuator reservoir.
  • the first on-actuator reservoir may include one or more on-actuator reservoirs.
  • the method of the invention for controlled dispensing of fluid into a droplet operations gap of a droplet actuator including a plurality of on-actuator reservoirs further includes use of an electrode arrangement corresponding to the plurality of on-actuator reservoirs, e.g., in which the electrode arrangement includes a plurality of sets of reservoir electrodes, in which each of the sets of the plurality of reservoir electrodes may be associated with one of the on-actuator reservoirs.
  • the electrode arrangement may include an array of sets of reservoir electrodes, e.g., the electrode arrangement may include 48 sets of reservoir electrodes, or the array of sets of reservoir electrodes may include a 4 x 12 array of sets reservoir electrodes.
  • the 4 x 12 array of sets reservoir electrodes may include four columns of sets of reservoir electrodes and twelve rows of sets of reservoir electrodes, in which each of the sets of reservoir electrodes may be at a location defined by its column location C0-C3 and its row location R0-R11.
  • the electrowetting-mediated dispensing of fluid from a first on-actuator reservoir may include dispensing of fluid using the first set of reservoir electrodes at a location defined by its column location C0-C3 and its row location R0-R1 1.
  • the electrowetting-mediated retention of fluid from one or more additional on-actuator reservoirs may include retaining the fluid from the one or more additional on-actuator reservoirs using one or more additional sets of reservoir electrodes at the same row location as the first set of reservoir electrodes, e.g., in which one reservoir electrode is activated in each of the one or more additional sets of reservoir electrodes at the same row location as the first set of reservoir electrodes.
  • Each of the sets of the plurality of reservoir electrodes may include three reservoir electrodes, in which the three reservoir electrodes comprise a reservoir electrode A, a reservoir electrode B, and a reservoir electrode C, e.g., in which reservoir electrode B may be activated in each of the one or more additional sets of reservoir electrodes at the same row location as the first set of reservoir electrodes.
  • the method of the invention for controlled dispensing of fluid into a droplet operations gap of a droplet actuator including a plurality of on-actuator reservoirs further includes dispensing of a sample fluid into the droplet operations gap.
  • the method of the invention for controlled dispensing of fluid into a droplet operations gap of a droplet actuator including a plurality of on-actuator reservoirs further includes the use of reservoir electrodes fluidly connected by an arrangement of droplet operations electrodes.
  • the invention provides a microfluidics system programmed to execute any of the methods described herein for controlled dispensing of fluid into a droplet operations gap of a droplet actuator including a plurality of on-actuator reservoirs.
  • the invention provides a microfluidics system programmed to execute any of the methods described herein for controlled loading of a droplet actuator with filler fluid.
  • the invention provides a microfluidics system programmed to execute any of the methods described herein, in which the method may include the use of any of the mechanisms disclosed herein.
  • the invention provides a storage medium including program code embodied in the medium for executing any of the methods disclosed herein for controlled dispensing of fluid into a droplet operations gap of a droplet actuator including a plurality of on- actuator reservoirs.
  • the invention provides a storage medium including program code embodied in the medium for executing any of the methods disclosed herein for controlled loading of a droplet actuator with filler fluid.
  • the invention provides a storage medium including program code embodied in the medium for executing any of the methods disclosed herein for controlled loading of a droplet actuator with filler fluid, in which the in which the method may include the use of any of the mechanisms disclosed herein, e.g., in which the mechanism may be coupled to a processor, or in which the processor executes program code embodied in a storage medium for executing the method for controlled loading of a droplet actuator with filler fluid.
  • the invention provides a microfluidics system including a droplet actuator including a plurality of on-actuator reservoirs, in which the droplet actuator may be coupled to a processor, and in which the processor executes program code embodied in a storage medium for executing any of the methods disclosed herein for controlled dispensing of fluid into a droplet operations gap of a droplet actuator including a plurality of on-actuator reservoirs.
  • Figure 1A illustrates a perspective view of an example of a filling fixture in use with a droplet actuator as per an aspect of an embodiment of the invention
  • Figure IB illustrates a cross-sectional view of the filling fixture of Figure 1A as per an aspect of an embodiment of the invention
  • Figure 2A illustrates a perspective view of another example of a filling fixture 1 A as per an aspect of an embodiment of the invention
  • Figure 2B illustrates a perspective view of the filling fixture of Figure 2A in use with a droplet actuator as per an aspect of an embodiment of the invention
  • Figures 3A and 3B illustrate perspective views of yet another example of a filling fixture as per an aspect of an embodiment of the invention
  • Figure 3C illustrates a perspective view of the filling fixture of Figures 3A and 3B in use with a droplet actuator as per an aspect of an embodiment of the invention
  • Figures 4A and 4B illustrate a perspective view and a cross-sectional view, respectively, of yet another example of a filling fixture as per an aspect of an embodiment of the invention
  • Figure 5A illustrates a perspective view of still another example of a filling fixture as per an aspect of an embodiment of the invention
  • Figure 5B illustrates a perspective view of the filling fixture of Figure 5A in use with a droplet actuator as per an aspect of an embodiment of the invention
  • Figures 6A and 6B illustrate side views of an example of a two-piece pipette for loading filler fluid into a droplet actuator as per an aspect of an embodiment of the invention
  • Figure 6C illustrates a perspective view of the two-piece pipette of Figures 6A and 6B iin use with a droplet actuator as per an aspect of an embodiment of the invention
  • Figure 7 illustrates a top view of an example of an electrode arrangement and a fluid reservoir dispensing protocol for improved control as per an aspect of an embodiment of the invention.
  • Figure 8 illustrates a functional block diagram of an example of a microfiuidics system that includes a droplet actuator as per an aspect of an embodiment of the invention. Definitions
  • Activate means affecting a change in the electrical state of the one or more electrodes which, in the presence of a droplet, results in a droplet operation.
  • Activation of an electrode can be accomplished using alternating or direct current. Any suitable voltage may be used.
  • an electrode may be activated using a voltage which is greater than about 150 V, or greater than about 200 V, or greater than about 250 V, or from about 275 V to about 1000 V, or about 300 V.
  • any suitable frequency may be employed.
  • an electrode may be activated using alternating current having a frequency from about 1 Hz to about 10 MHz, or from about 10 Hz to about 60 Hz, or from about 20 Hz to about 40 Hz, or about 30 Hz.
  • Bead with respect to beads on a droplet actuator, means any bead or particle that is capable of interacting with a droplet on or in proximity with a droplet actuator.
  • Beads may be any of a wide variety of shapes, such as spherical, generally spherical, egg shaped, disc shaped, cubical, amorphous and other three dimensional shapes.
  • the bead may, for example, be capable of being subjected to a droplet operation in a droplet on a droplet actuator or otherwise configured with respect to a droplet actuator in a manner which permits a droplet on the droplet actuator to be brought into contact with the bead on the droplet actuator and/or off the droplet actuator.
  • Beads may be provided in a droplet, in a droplet operations gap, or on a droplet operations surface. Beads may be provided in a reservoir that is external to a droplet operations gap or situated apart from a droplet operations surface, and the reservoir may be associated with a flow path that permits a droplet including the beads to be brought into a droplet operations gap or into contact with a droplet operations surface. Beads may be manufactured using a wide variety of materials, including for example, resins, and polymers. The beads may be any suitable size, including for example, microbeads, microparticles, nanobeads and nanoparticles. In some cases, beads are magnetically responsive; in other cases beads are not significantly magnetically responsive.
  • the magnetically responsive material may constitute substantially all of a bead, a portion of a bead, or only one component of a bead.
  • the remainder of the bead may include, among other things, polymeric material, coatings, and moieties which permit attachment of an assay reagent.
  • suitable beads include flow cytometry microbeads, polystyrene microparticles and nanoparticles, functionalized polystyrene microparticles and nanoparticles, coated polystyrene microparticles and nanoparticles, silica microbeads, fluorescent microspheres and nanospheres, functionalized fluorescent microspheres and nanospheres, coated fluorescent microspheres and nanospheres, color dyed microparticles and nanoparticles, magnetic microparticles and nanoparticles, superparamagnetic microparticles and nanoparticles (e.g., DYNABEADS® particles, available from Invitrogen Group, Carlsbad, CA), fluorescent microparticles and nanoparticles, coated magnetic microparticles and nanoparticles, ferromagnetic microparticles and nanoparticles, coated ferromagnetic microparticles and nanoparticles, and those described in U.S.
  • DYNABEADS® particles available from Invitrogen Group, Carlsbad,
  • Beads may be pre-coupled with a biomolecule or other substance that is able to bind to and form a complex with a biomolecule. Beads may be pre-coupled with an antibody, protein or antigen, DNA/RNA probe or any other molecule with an affinity for a desired target.
  • droplet actuator techniques for immobilizing magnetically responsive beads and/or non-magnetically responsive beads and/or conducting droplet operations protocols using beads are described in U.S. Patent Application No. 1 1/639,566, entitled “Droplet-Based Particle Sorting," filed on December 15, 2006; U.S. Patent Application No. 61/039,183, entitled “Multiplexing Bead Detection in a Single Droplet,” filed on March 25, 2008; U.S.
  • Patent Application No. 61/047,789 entitled “Droplet Actuator Devices and Droplet Operations Using Beads," filed on April 25, 2008
  • U.S. Patent Application No. 61/086, 183 entitled “Droplet Actuator Devices and Methods for Manipulating Beads,” filed on August 5, 2008
  • International Patent Application No. PCT/US2008/053545 entitled “Droplet Actuator Devices and Methods Employing Magnetic Beads,” filed on February 11, 2008
  • International Patent Application No. PCT/US2008/058018 entitled “Bead-based Multiplexed Analytical Methods and Instrumentation,” filed on March 24, 2008
  • Droplet means a volume of liquid on a droplet actuator.
  • a droplet is at least partially bounded by a filler fluid.
  • a droplet may be completely surrounded by a filler fluid or may be bounded by filler fluid and one or more surfaces of the droplet actuator.
  • a droplet may be bounded by filler fluid, one or more surfaces of the droplet actuator, and/or the atmosphere.
  • a droplet may be bounded by filler fluid and the atmosphere.
  • Droplets may, for example, be aqueous or non-aqueous or may be mixtures or emulsions including aqueous and non-aqueous components.
  • Droplets may take a wide variety of shapes; nonlimiting examples include generally disc shaped, slug shaped, truncated sphere, ellipsoid, spherical, partially compressed sphere, hemispherical, ovoid, cylindrical, combinations of such shapes, and various shapes formed during droplet operations, such as merging or splitting or formed as a result of contact of such shapes with one or more surfaces of a droplet actuator.
  • droplet fluids that may be subjected to droplet operations using the approach of the invention, see International Patent Application No. PCT/US 06/47486, entitled, "Droplet- Based Biochemistry," filed on December 1 1, 2006.
  • a droplet may include a biological sample, such as whole blood, lymphatic fluid, serum, plasma, sweat, tear, saliva, sputum, cerebrospinal fluid, amniotic fluid, seminal fluid, vaginal excretion, serous fluid, synovial fluid, pericardial fluid, peritoneal fluid, pleural fluid, transudates, exudates, cystic fluid, bile, urine, gastric fluid, intestinal fluid, fecal samples, liquids containing single or multiple cells, liquids containing organelles, fluidized tissues, fluidized organisms, liquids containing multi- celled organisms, biological swabs and biological washes.
  • a biological sample such as whole blood, lymphatic fluid, serum, plasma, sweat, tear, saliva, sputum, cerebrospinal fluid, amniotic fluid, seminal fluid, vaginal excretion, serous fluid, synovial fluid, pericardial fluid, peritoneal fluid, pleural fluid, transudates, exu
  • a droplet may include a reagent, such as water, deionized water, saline solutions, acidic solutions, basic solutions, detergent solutions and/or buffers.
  • reagents such as a reagent for a biochemical protocol, such as a nucleic acid amplification protocol, an affinity- based assay protocol, an enzymatic assay protocol, a sequencing protocol, and/or a protocol for analyses of biological fluids.
  • a droplet may include one or more beads.
  • Droplet Actuator means a device for manipulating droplets.
  • droplet actuators see Pamula et al., U.S. Patent 6,91 1, 132, entitled “Apparatus for Manipulating Droplets by Electrowetting-Based Techniques,” issued on June 28, 2005; Pamula et al., U.S. Patent Application No. 1 1/343,284, entitled “Apparatuses and Methods for Manipulating Droplets on a Printed Circuit Board,” filed on filed on January 30, 2006; Pollack et al., International Patent Application No. PCT/US2006/047486, entitled “Droplet-Based Biochemistry,” filed on December 1 1, 2006; Shenderov, U.S.
  • Patent 7,547,380 entitled “Droplet Transportation Devices and Methods Having a Fluid Surface,” issued on June 16, 2009; Sterling et al., U.S. Patent 7, 163,612, entitled “Method, Apparatus and Article for Microfluidic Control via Electrowetting, for Chemical, Biochemical and Biological Assays and the Like,” issued on January 16, 2007; Becker and Gascoyne et al., U.S. Patent Nos. 7,641,779, entitled “Method and Apparatus for Programmable fluidic Processing,” issued on January 5, 2010, and 6,977,033, entitled “Method and Apparatus for Programmable fluidic Processing,” issued on December 20, 2005; Deere et al., U.S.
  • Patent 7,328,979 entitled “System for Manipulation of a Body of Fluid,” issued on February 12, 2008; Yamakawa et al., U.S. Patent Pub. No. 20060039823, entitled “Chemical Analysis Apparatus,” published on February 23, 2006; Wu, International Patent Pub. No. WO/2009/003184, entitled “Digital Microfluidics Based Apparatus for Heat-exchanging Chemical Processes,” published on December 31, 2008; Fouillet et al., U.S. Patent Pub. No. 20090192044, entitled “Electrode Addressing Method,” published on July 30, 2009; Fouillet et al., U.S.
  • Patent 7,052,244 entitled “Device for Displacement of Small Liquid Volumes Along a Micro-catenary Line by Electrostatic Forces,” issued on May 30, 2006; Marchand et al., U.S. Patent Pub. No. 20080124252, entitled “Droplet Microreactor,” published on May 29, 2008; Adachi et al., U.S. Patent Pub. No. 20090321262, entitled “Liquid Transfer Device,” published on December 31, 2009; Roux et al., U.S. Patent Pub. No.
  • Certain droplet actuators will include one or more substrates arranged with a droplet operations gap therebetween and electrodes associated with (e.g., layered on, attached to, and/or embedded in) the one or more substrates and arranged to conduct one or more droplet operations.
  • certain droplet actuators will include a base (or bottom) substrate, droplet operations electrodes associated with the substrate, one or more dielectric layers atop the substrate and/or electrodes, and optionally one or more hydrophobic layers atop the substrate, dielectric layers and/or the electrodes forming a droplet operations surface.
  • a top substrate may also be provided, which is separated from the droplet operations surface by a gap, commonly referred to as a droplet operations gap.
  • a droplet operations gap commonly referred to as a droplet operations gap.
  • a ground or reference electrode may be associated with the top substrate facing the gap, the bottom substrate facing the gap, in the gap.
  • electrical contacts for coupling the electrodes to a droplet actuator instrument for controlling or monitoring the electrodes may be associated with one or both plates.
  • electrodes on one substrate are electrically coupled to the other substrate so that only one substrate is in contact with the droplet actuator.
  • a conductive material e.g., an epoxy, such as MASTER BONDTM Polymer System EP79, available from Master Bond, Inc., Hackensack, NJ
  • a conductive material provides the electrical connection between electrodes on one substrate and electrical paths on the other substrates, e.g., a ground electrode on a top substrate may be coupled to an electrical path on a bottom substrate by such a conductive material.
  • a spacer may be provided between the substrates to determine the height of the gap therebetween and define dispensing reservoirs.
  • the spacer height may, for example, be from about 5 ⁇ to about 600 ⁇ , or about 100 ⁇ to about 400 ⁇ , or about 200 ⁇ to about 350 ⁇ , or about 250 ⁇ to about 300 ⁇ , or about 275 ⁇ .
  • the spacer may, for example, be formed of a layer of projections form the top or bottom substrates, and/or a material inserted between the top and bottom substrates.
  • One or more openings may be provided in the one or more substrates for forming a fluid path through which liquid may be delivered into the droplet operations gap.
  • the one or more openings may in some cases be aligned for interaction with one or more electrodes, e.g., aligned such that liquid flowed through the opening will come into sufficient proximity with one or more droplet operations electrodes to permit a droplet operation to be effected by the droplet operations electrodes using the liquid.
  • the base (or bottom) and top substrates may in some cases be formed as one integral component.
  • One or more reference electrodes may be provided on the base (or bottom) and/or top substrates and/or in the gap. Examples of reference electrode arrangements are provided in the above referenced patents and patent applications.
  • the manipulation of droplets by a droplet actuator may be electrode mediated, e.g., electrowetting-mediated or dielectrophoresis mediated or Coulombic force mediated.
  • electrode mediated e.g., electrowetting-mediated or dielectrophoresis mediated or Coulombic force mediated.
  • other techniques for controlling droplet operations include using devices that induce hydrodynamic fluidic pressure, such as those that operate on the basis of mechanical principles (e.g. external syringe pumps, pneumatic membrane pumps, vibrating membrane pumps, vacuum devices, centrifugal forces, piezoelectric/ultrasonic pumps and acoustic forces); electrical or magnetic principles (e.g.
  • thermodynamic principles e.g. gas bubble generation/phase- change-induced volume expansion
  • other kinds of surface-wetting principles e.g. electrowetting, and optoelectrowetting, as well as chemically, thermally, structurally and radioactively induced surface-tension gradients
  • gravity e.g., capillary action
  • electrostatic forces e.g., electroosmotic flow
  • centrifugal flow substrate disposed on a compact disc and rotated
  • magnetic forces e.g., oscillating ions causes flow
  • magnetohydrodynamic forces and vacuum or pressure differential.
  • combinations of two or more of the foregoing techniques may be employed to conduct a droplet operation in a droplet actuator of the invention.
  • one or more of the foregoing may be used to deliver liquid into a droplet operations gap, e.g., from a reservoir in another device or from an external reservoir of the droplet actuator (e.g., a reservoir associated with a droplet actuator substrate and a flow path from the reservoir into the droplet operations gap).
  • Droplet operations surfaces of certain droplet actuators of the invention may be made from hydrophobic materials or may be coated or treated to make them hydrophobic.
  • some portion or all of the droplet operations surfaces may be derivatized with low surface-energy materials or chemistries, e.g., by deposition or using in situ synthesis using compounds such as poly- or per-fluorinated compounds in solution or polymerizable monomers.
  • Examples include TEFLON® AF (available from DuPont, Wilmington, DE), members of the cytop family of materials, coatings in the FLUOROPEL® family of hydrophobic and superhydrophobic coatings (available from Cytonix Corporation, Beltsville, MD), silane coatings, fluorosilane coatings, hydrophobic phosphonate derivatives (e.g.., those sold by Aculon, Inc), and NOVECTM electronic coatings (available from 3M Company, St. Paul, MN), other fluorinated monomers for plasma-enhanced chemical vapor deposition (PECVD), and organosiloxane (e.g., SiOC) for PECVD.
  • PECVD plasma-enhanced chemical vapor deposition
  • organosiloxane e.g., SiOC
  • the droplet operations surface may include a hydrophobic coating having a thickness ranging from about 10 nm to about 1 ,000 nm.
  • the top substrate of the droplet actuator includes an electrically conducting organic polymer, which is then coated with a hydrophobic coating or otherwise treated to make the droplet operations surface hydrophobic.
  • the electrically conducting organic polymer that is deposited onto a plastic substrate may be poly(3,4-ethylenedioxythiophene) poly(styrenesulfonate) (PEDOT:PSS).
  • PEDOT:PSS poly(3,4-ethylenedioxythiophene) poly(styrenesulfonate)
  • Other examples of electrically conducting organic polymers and alternative conductive layers are described in Pollack et al., International Patent Application No.
  • One or both substrates may be fabricated using a printed circuit board (PCB), glass, indium tin oxide (ITO)-coated glass, and/or semiconductor materials as the substrate.
  • the ITO coating is preferably a thickness in the range of about 20 to about 200 nm, preferably about 50 to about 150 nm, or about 75 to about 125 nm, or about 100 nm.
  • the top and/or bottom substrate includes a PCB substrate that is coated with a dielectric, such as a polyimide dielectric, which may in some cases also be coated or otherwise treated to make the droplet operations surface hydrophobic.
  • a dielectric such as a polyimide dielectric
  • the substrate includes a PCB
  • the following materials are examples of suitable materials: MITSUITM BN-300 (available from MITSUI Chemicals America, Inc., San Jose CA); ARLONTM 1 IN (available from Arlon, Inc, Santa Ana, CA).; NELCO® N4000-6 and N5000-30/32 (available from Park Electrochemical Corp., Melville, NY); ISOLATM FR406 (available from Isola Group, Chandler, AZ), especially IS620; fluoropolymer family (suitable for fluorescence detection since it has low background fluorescence); polyimide family; polyester; polyethylene naphthalate; polycarbonate; polyetheretherketone; liquid crystal polymer; cyclo-olefm copolymer (
  • Various materials are also suitable for use as the dielectric component of the substrate. Examples include: vapor deposited dielectric, such as PARYLENETM C (especially on glass), PARYLENETM N, and PARYLENETM HT (for high temperature, ⁇ 300°C) (available from Parylene Coating Services, Inc., Katy, TX); TEFLON® AF coatings; cytop; soldermasks, such as liquid photoimageable soldermasks (e.g., on PCB) like TAIYOTM PSR4000 series, TAIYOTM PSR and AUS series (available from Taiyo America, Inc.
  • vapor deposited dielectric such as PARYLENETM C (especially on glass), PARYLENETM N, and PARYLENETM HT (for high temperature, ⁇ 300°C) (available from Parylene Coating Services, Inc., Katy, TX); TEFLON® AF coatings; cytop; soldermasks, such as liquid photoimageable soldermas
  • Droplet transport voltage and frequency may be selected for performance with reagents used in specific assay protocols.
  • Design parameters may be varied, e.g., number and placement of on-actuator reservoirs, number of independent electrode connections, size (volume) of different reservoirs, placement of magnets/bead washing zones, electrode size, inter-electrode pitch, and gap height (between top and bottom substrates) may be varied for use with specific reagents, protocols, droplet volumes, etc.
  • a substrate of the invention may derivatized with low surface- energy materials or chemistries, e.g., using deposition or in situ synthesis using poly- or per-fluorinated compounds in solution or polymerizable monomers.
  • the droplet operations surface may be coated with a substance for reducing background noise, such as background fluorescence from a PCB substrate.
  • the noise-reducing coating may include a black matrix resin, such as the black matrix resins available from Toray industries, Inc., Japan.
  • Electrodes of a droplet actuator are typically controlled by a controller or a processor, which is itself provided as part of a system, which may include processing functions as well as data and software storage and input and output capabilities.
  • Reagents may be provided on the droplet actuator in the droplet operations gap or in a reservoir fluidly coupled to the droplet operations gap.
  • the reagents may be in liquid form, e.g., droplets, or they may be provided in a reconstitutable form in the droplet operations gap or in a reservoir fluidly coupled to the droplet operations gap.
  • Reconstitutable reagents may typically be combined with liquids for reconstitution.
  • An example of reconstitutable reagents suitable for use with the invention includes those described in Meathrel, et al., U.S. Patent 7,727,466, entitled “Disintegratable films for diagnostic devices," granted on June 1, 2010.
  • Droplet operation means any manipulation of a droplet on a droplet actuator.
  • a droplet operation may, for example, include: loading a droplet into the droplet actuator; dispensing one or more droplets from a source droplet; splitting, separating or dividing a droplet into two or more droplets; transporting a droplet from one location to another in any direction; merging or combining two or more droplets into a single droplet; diluting a droplet; mixing a droplet; agitating a droplet; deforming a droplet; retaining a droplet in position; incubating a droplet; heating a droplet; vaporizing a droplet; cooling a droplet; disposing of a droplet; transporting a droplet out of a droplet actuator; other droplet operations described herein; and/or any combination of the foregoing.
  • merge “merge,” “merging,” “combine,” “combining” and the like are used to describe the creation of one droplet from two or more droplets. It should be understood that when such a term is used in reference to two or more droplets, any combination of droplet operations that are sufficient to result in the combination of the two or more droplets into one droplet may be used. For example, “merging droplet A with droplet B,” can be achieved by transporting droplet A into contact with a stationary droplet B, transporting droplet B into contact with a stationary droplet A, or transporting droplets A and B into contact with each other.
  • splitting is not intended to imply any particular outcome with respect to volume of the resulting droplets (i.e., the volume of the resulting droplets can be the same or different) or number of resulting droplets (the number of resulting droplets may be 2, 3, 4, 5 or more).
  • mixing refers to droplet operations which result in more homogenous distribution of one or more components within a droplet. Examples of “loading” droplet operations include microdialysis loading, pressure assisted loading, robotic loading, passive loading, and pipette loading. Droplet operations may be electrode-mediated. In some cases, droplet operations are further facilitated by the use of hydrophilic and/or hydrophobic regions on surfaces and/or by physical obstacles.
  • Impedance or capacitance sensing or imaging techniques may sometimes be used to determine or confirm the outcome of a droplet operation. Examples of such techniques are described in Sturmer et al., U.S. Patent Application Publication No. US20100194408, entitled “Capacitance Detection in a Droplet Actuator,” published on Aug. 5, 2010, the entire disclosure of which is incorporated herein by reference. Generally speaking, the sensing or imaging techniques may be used to confirm the presence or absence of a droplet at a specific electrode.
  • the presence of a dispensed droplet at the destination electrode following a droplet dispensing operation confirms that the droplet dispensing operation was effective.
  • the presence of a droplet at a detection spot at an appropriate step in an assay protocol may confirm that a previous set of droplet operations has successfully produced a droplet for detection.
  • Droplet transport time can be quite fast. For example, in various embodiments, transport of a droplet from one electrode to the next may exceed about 1 sec, or about 0.1 sec, or about 0.01 sec, or about 0.001 sec.
  • the electrode is operated in AC mode but is switched to DC mode for imaging.
  • droplet operations for the footprint area of droplet are similar to electrowetting area; in other words, lx-, 2x- 3x-droplets are usefully controlled operated using 1, 2, and 3 electrodes, respectively. If the droplet footprint is greater than the number of electrodes available for conducting a droplet operation at a given time, the difference between the droplet size and the number of electrodes should typically not be greater than 1 ; in other words, a 2x droplet is usefully controlled using 1 electrode and a 3x droplet is usefully controlled using 2 electrodes. When droplets include beads, it is useful for droplet size to be equal to the number of electrodes controlling the droplet, e.g., transporting the droplet.
  • Filler fluid means a fluid associated with a droplet operations substrate of a droplet actuator, which fluid is sufficiently immiscible with a droplet phase to render the droplet phase subject to electrode-mediated droplet operations.
  • the droplet operations gap of a droplet actuator is typically filled with a filler fluid.
  • the filler fluid may, for example, be or include a low- viscosity oil, such as silicone oil or hexadecane filler fluid.
  • the filler fluid may be or include a halogenated oil, such as a fluorinated or perfluorinated oil.
  • the filler fluid may fill the entire gap of the droplet actuator or may coat one or more surfaces of the droplet actuator. Filler fluids may be conductive or non-conductive.
  • Filler fluids may be selected to improve droplet operations and/or reduce loss of reagent or target substances from droplets, improve formation of microdroplets, reduce cross contamination between droplets, reduce contamination of droplet actuator surfaces, reduce degradation of droplet actuator materials, etc.
  • filler fluids may be selected for compatibility with droplet actuator materials.
  • fluorinated filler fluids may be usefully employed with fluorinated surface coatings.
  • Fluorinated filler fluids are useful to reduce loss of lipophilic compounds, such as umbelliferone substrates like 6- hexadecanoylamido-4-methylumbelliferone substrates (e.g., for use in Krabbe, Niemann-Pick, or other assays); other umbelliferone substrates are described in U.S. Patent Pub. No. 201 101 18132, published on May 19, 201 1, the entire disclosure of which is incorporated herein by reference.
  • filler fluids are based on kinematic viscosity ( ⁇ 7 cSt is preferred, but not required), and on boiling point (> 150 °C is preferred, but not required, for use in DNA/RNA-based applications (PCR, etc.)).
  • Filler fluids may, for example, be doped with surfactants or other additives.
  • additives may be selected to improve droplet operations and/or reduce loss of reagent or target substances from droplets, formation of microdroplets, cross contamination between droplets, contamination of droplet actuator surfaces, degradation of droplet actuator materials, etc.
  • Composition of the filler fluid may be selected for performance with reagents used in the specific assay protocols and effective interaction or non-interaction with droplet actuator materials.
  • filler fluids and filler fluid formulations suitable for use with the invention are provided in Srinivasan et al, International Patent Pub. Nos. WO/2010/027894, entitled “Droplet Actuators, Modified Fluids and Methods,” published on March 1 1, 2010, and WO/2009/021 173, entitled “Use of Additives for Enhancing Droplet Operations,” published on February 12, 2009; Sista et al., International Patent Pub. No.
  • Fluorinated oils may in some cases be doped with fluorinated surfactants, e.g., Zonyl FSO-100 (Sigma-Aldrich) and/or others.
  • a droplet actuator system of the invention may include on-cartridge reservoirs and/or off- cartridge reservoirs.
  • On-cartridge reservoirs may be (1) on-actuator reservoirs, which are reservoirs in the droplet operations gap or on the droplet operations surface; (2) off-actuator reservoirs, which are reservoirs on the droplet actuator cartridge, but outside the droplet operations gap, and not in contact with the droplet operations surface; or (3) hybrid reservoirs which have on-actuator regions and off-actuator regions.
  • An example of an off-actuator reservoir is a reservoir in the top substrate.
  • An off-actuator reservoir is typically in fluid communication with an opening or flow path arranged for flowing liquid from the off-actuator reservoir into the droplet operations gap, such as into an on-actuator reservoir.
  • An off-cartridge reservoir may be a reservoir that is not part of the droplet actuator cartridge at all, but which flows liquid to some portion of the droplet actuator cartridge.
  • an off-cartridge reservoir may be part of a system or docking station to which the droplet actuator cartridge is coupled during operation.
  • an off-cartridge reservoir may be a reagent storage container or syringe which is used to force fluid into an on-cartridge reservoir or into a droplet operations gap.
  • a system using an off-cartridge reservoir will typically include a fluid passage means whereby liquid may be transferred from the off-cartridge reservoir into an on-cartridge reservoir or into a droplet operations gap.
  • top bottom
  • over under
  • under on
  • the terms “top,” “bottom,” “over,” “under,” and “on” are used throughout the description with reference to the relative positions of components of the droplet actuator, such as relative positions of top and bottom substrates of the droplet actuator. It will be appreciated that the droplet actuator is functional regardless of its orientation in space.
  • a liquid in any form e.g., a droplet or a continuous body, whether moving or stationary
  • a liquid in any form e.g., a droplet or a continuous body, whether moving or stationary
  • such liquid could be either in direct contact with the electrode/array/matrix/surface, or could be in contact with one or more layers or films that are interposed between the liquid and the electrode/array/matrix/surface.
  • filler fluid can be considered as a film between such liquid and the electrode/array/matrix/surface.
  • a droplet When a droplet is described as being “on” or “loaded on” a droplet actuator, it should be understood that the droplet is arranged on the droplet actuator in a manner which facilitates using the droplet actuator to conduct one or more droplet operations on the droplet, the droplet is arranged on the droplet actuator in a manner which facilitates sensing of a property of or a signal from the droplet, and/or the droplet has been subjected to a droplet operation on the droplet actuator.
  • the present invention is directed to mechanisms for and methods of loading a droplet actuator with filler fluid.
  • the mechanisms and methods are useful for loading filler fluid in a droplet actuator in a manner that is rapid enough for practical use while at the same time sufficiently slow to reduce, preferably entirely avoid, the trapping of air in the droplet operations gap during the loading process.
  • Embodiments of the invention include filling fixtures that can be mechanically and fluidly coupled to the filler fluid loading port of a droplet actuator, wherein the filling fixtures provide funnel-like functions for loading filler fluid by gravity feed. Certain embodiments of the filling fixtures allow the filler fluid loading process to be highly visually observable.
  • the present invention is also directed to a fluid reservoir dispensing protocol for improved control.
  • An aspect of an embodiment of the invention may include a filling fixture having an outlet physically connected to an outlet shroud including where the outlet is capable of communicating fluid from the filling fixture to a droplet actuator.
  • An aspect of an embodiment of the invention may include a reservoir in fluid communication with the outlet.
  • An aspect of an embodiment of the invention may include a filling fixture having an opening (1 12).
  • a further aspect of an embodiment of the invention may include a filling fixture having an opening (1 12), including wherein the opening (1 12) has a diameter d3 where d3 may be from about .02 inches to about .05 inches.
  • An additional aspect of an embodiment of the invention may include a filling fixture having an opening (1 12) having a diameter d3 from about .04 inches to about .05 inches.
  • An additional aspect of an embodiment of the invention may include a filling fixture having an outlet (1 12) having a diameter d3 from about .04 inches to about .05 inches.
  • a further aspect of an embodiment of the invention may include a filling fixture having an opening (1 12) having a diameter d3 of about .047 inches.
  • An additional aspect of an embodiment of the invention may include a droplet actuator having a filling fixture (1 10) physically connected to a corner of a droplet actuator.
  • Still another aspect of an embodiment of the invention may include a droplet actuator having a filling fixture (1 10) physically connected to a droplet actuator at a corner of the droplet actuator opposite the electrical contact points.
  • An additional aspect of an embodiment of the invention may include a droplet actuator having a filling fixture (1 10) physically connected to a droplet actuator at a corner of the droplet actuator having electrical contact points.
  • Figure 1A illustrates a perspective view of an example of a filling fixture 100 in use with a droplet actuator 150.
  • Droplet actuator 150 includes a bottom substrate 152 and a top substrate 154 that are separated by a droplet operations gap.
  • Droplet actuator 150 includes a plurality of on-actuator reservoirs for processing, for example, sample fluid and reagent fluid.
  • Top substrate 154 includes a plurality of loading ports 156 for loading fluid into the on-actuator reservoirs.
  • a port is an entrance/exit (opening) to the droplet operations gap. Liquid may flow through the port into and/or from any portion of the droplet operations gap.
  • Each port provides a fluid path through top substrate 154 to the droplet operations gap between bottom substrate 152 and top substrate 154.
  • top substrate 154 includes a filler fluid loading port (not visible beneath the filling fixture 100) for loading filler fluid into the droplet operations gap.
  • the filler fluid is, for example, low- viscosity oil, such as silicone oil or hexadecane filler fluid.
  • the top substrate and the bottom substrate separated by the droplet operations gap may comprise a single integral substrate.
  • a single integral substrate may be folded over on itself to form a top portion and a bottom portion comprising the top substrate and ihe botiom substrate, respectively.
  • the top substrate and the bottom substrate separated by the droplet operations gap may also be formed, for example, by carving a gap out of a single integral substrate or by creating a gap in a single integral substrate using semiconductor etching techniques.
  • the filler fluid loading port includes, for example, a cylindrical element (or feature) around the opening in top substrate 154 and that protrudes outwardly from the top substrate 154 (i.e., away from the droplet operations gap).
  • FIG. 1A shows the filling fixture 100 mechanically and fluidly coupled to the filler fluid loading port of top substrate 154.
  • Filling fixture 100 includes a cylinder- or dish-shaped filling fixture fluid reservoir 1 10 for holding a volume of filler fluid (not shown).
  • Filling fixture fluid reservoir 1 10 includes an outlet 1 12, which is an opening in the floor of filling fixture fluid reservoir 1 10.
  • Filler fluid may flow through outlet 1 12 by gravity and exit filling fixture fluid reservoir 1 10.
  • Filling fixture 100 further includes an outlet shroud 1 14 that is positioned around outlet 1 12 on the underside of filling fixture fluid reservoir 1 10.
  • Outlet shroud 1 14 is designed to be mechanically coupled to the filler fluid loading port in top substrate 154.
  • outlet shroud 1 14 of filling fixture 100 is designed to be press- fitted onto the filler fluid loading port in top substrate 154.
  • outlet 1 12 of filling fixture 100 is aligned with the filler fluid loading port in top substrate 154.
  • outlet 1 12 is fluidly coupled to the filler fluid loading port in top substrate 154.
  • opening 1 12 and outlet shroud 1 14 are substantially centered with respect to filling fixture fluid reservoir 1 10.
  • Figure IB illustrates a cross-sectional view of filling fixture 100 in use with droplet actuator 150.
  • outlet 112 is substantially centered with respect to filling fixture fluid reservoir 1 10.
  • Figure IB shows a filler fluid loading port 158 in top substrate 154.
  • filler fluid loading port 158 is a cylindrical element around an opening in top substrate 154.
  • the opening in top substrate 154 has a diameter dl .
  • the diameter dl is also the inside diameter of filler fluid loading port 158.
  • filler fluid loading port 158 has an outside diameter d2.
  • the diameter dl is, for example, from about .01 13 inches to about .0913 inches, or from about .0413 inches to about .0613 inches, or about .0513 inches.
  • the diameter d2 is, for example, from about .09 inches to about .13 inches, or about .1 1 inches.
  • opening 112 has a diameter d3.
  • the diameter d3 is, for example, from about 0.001 inches to about 0.1 inches, from about 0.01 inches to about 0.06 inches, from about 0.02 inches to about 0.05 inches, from about .04 inches to about .05 inches, or about 0.047 inches.
  • Filling fixture fluid reservoir 1 10 of filling fixture 100 is designed to hold a certain volume VI of filler fluid.
  • filling fixture fluid reservoir 1 10 may hold from about 1 ml to about 1.5 ml, or about 1.26 ml of filler fluid.
  • filling fixture fluid reservoir 1 10 has a height h, which is, for example, from about 7.5 mm to about 8.5 mm, or about 8 mm.
  • the floor of filling fixture fluid reservoir 1 10 can be substantially flat or be slightly counter-sunk leading to opening 112, as shown in Figure IB.
  • the inside diameter of outlet shroud 1 14 of filling fixture 100 is at least slightly greater than the outside diameter d2 of filler fluid loading port 158.
  • the inside diameter of outlet shroud 1 14 is sized to allow outlet shroud 1 14 to be press fitted onto filler fluid loading port 158.
  • a user press fits outlet shroud 1 14 of filling fixture 100 onto filler fluid loading port 158 in top substrate 154, thereby mechanically and fluidly coupling filling fixture 100 to droplet actuator 150.
  • filling fixture 100 is fitted onto filler fluid loading port 158 like a cap.
  • the user then fills the filling fixture fluid reservoir 1 10 of filling fixture 100 with a volume of filler fluid.
  • the filler fluid flows by gravity through outlet 1 12 of filling fixture 100, through filler fluid loading port 158, and into the droplet operations gap of droplet actuator 150.
  • Loading the filler fluid too rapidly into a droplet actuator can cause air to be trapped in the droplet operations gap of the droplet actuator. Additionally, loading the filler fluid too rapidly can cause the loss of filler fluid through other openings (e.g., sample or reagent loading ports) in the top substrate of a droplet actuator.
  • the flow of filler fluid into the droplet operations gap of droplet actuator 150 is provided in a controlled fashion that is rapid enough for practical use, while at the same time sufficiently slow to (1) reduce, preferably entirely eliminate, trapping of air in the droplet operations gap and (2) reduce, preferably entirely eliminate, the loss of filler fluid through openings, such as loading ports 156, in top substrate 154.
  • Filling fixture 100 is not limited to the shape and features shown in Figures 1A and IB. Namely, filling fixture 100 can be implemented in a variety of shapes and with a variety of features, examples of which are described with reference to Figures 2A through 6C.
  • FIG 2A illustrates a perspective view of another example of filling fixture 100.
  • outlet 1 12 is positioned offset from center.
  • outlet 1 12 and its corresponding outlet shroud 114 are positioned substantially offset from the center of filling fixture fluid reservoir 1 10.
  • the offset from center outlet 1 12 and outlet shroud 1 14 allows the majority of filling fixture fluid reservoir 1 10 to be positioned off the edge of droplet actuator 150 when in use.
  • the user's view of filler fluid loading into the droplet operations gap of droplet actuator 150 is largely unobstructed by filling fixture fluid reservoir 1 10.
  • FIGs 3A and 3B illustrate perspective views of yet another example of filling fixture 100.
  • filling fixture fluid reservoir 1 10 of filling fixture 100 is kidney- or piano-shaped.
  • Figure 3 A shows the filling fixture fluid reservoir 1 10-side of filling fixture 100
  • Figure 3B shows the outlet shroud 1 14-side of filling fixture 100.
  • Outlet 112 and outlet shroud 1 14 are positioned to one side of the kidney- or piano-shaped filling fixture fluid reservoir 1 10 in order to allow the user's unobstructed view of the filler fluid loading process, which is shown in Figure 3C.
  • Figure 3C illustrates a perspective view of the filling fixture 100 of Figures 3 A and 3B in use with droplet actuator 150.
  • outlet shroud 1 14 of filling fixture 100 is press fitted onto filler fluid loading port 158 of droplet actuator 150.
  • the majority of filling fixture fluid reservoir 1 10 is positioned off the edge of droplet actuator 150.
  • the user's view of filler fluid loading into the droplet operations gap of droplet actuator 150 is largely unobstructed by the kidney- or piano-shaped filling fixture fluid reservoir 1 10.
  • Figures 4A and 4B illustrate a perspective view and a cross-sectional view, respectively, of yet another example of filling fixture 100.
  • filling fixture 100 is substantially the same as filling fixture 100 of Figures 1A and IB, except that outlet shroud 1 14 is omitted and replaced with an outlet tip 120.
  • Outlet tip 120 is a tapered, pipette-like tip that is designed to be fitted into the opening of filler fluid loading port 158, as shown in Figure 4B. While Figures 4A and 4B show outlet 1 12 and outlet tip 120 substantially centered with respect to filling fixture fluid reservoir 1 10, in other implementations outlet 1 12 and outlet tip 120 can be positioned off center to ensure a largely unobstructed view of the filler fluid loading process.
  • FIG. 5A illustrates a perspective view of still another example of filling fixture 100.
  • Figure 5B illustrates a perspective view of filling fixture 100 of Figure 5A in use with droplet actuator 150.
  • filling fixture fluid reservoir 1 10 of filling fixture 100 is trough-shaped and includes outlet tip 120. While Figures 5A and 5B show outlet 1 12 and outlet tip 120 substantially centered with respect to the trough-shaped filling fixture fluid reservoir 1 10, in other implementations outlet 1 12 and outlet tip 120 can be positioned off center to ensure a largely unobstructed view of the filler fluid loading process. Additionally, instead of outlet tip 120, outlet shroud 1 14 may be provided at outlet 1 12 of the trough-shaped filling fixture fluid reservoir 1 10.
  • the filling fixture fluid reservoir 1 10 of filling fixtures 100 can be designed with any shape and to hold any volume of filler fluid.
  • outlet 1 12 can be placed in any location with respect to filling fixture fluid reservoir 1 10.
  • the shape of filling fixture fluid reservoir 1 10 and the location of outlet 1 12 ensure the user's unobstructed view of the filler fluid loading process when in use.
  • Figures 6A and 6B illustrate side views of an example of a two-piece pipette 600 for loading filler fluid into a droplet actuator, such as into droplet actuator 150.
  • Figure 6A shows two-piece pipette 600 in an unassembled state
  • Figure 6B shows two-piece pipette 600 in an assembled state.
  • two-piece pipette 600 includes a first pipette tip 610 that is fitted into a second pipette tip 620.
  • First pipette tip 610 is a hollow tapered tube that is open at each end. Namely, first pipette tip 610 includes an inlet 612 and an outlet 614, wherein inlet 612 has a larger diameter than outlet 614.
  • second pipette tip 620 is a hollow tapered tube that is open at each end. Namely, second pipette tip 620 includes an inlet 622 and an outlet 624, wherein inlet 622 has a larger diameter than outlet 624.
  • First pipette tip 610 and second pipette tip 620 may be formed, for example, of molded plastic. Outlet 614 of first pipette tip 610 is sized to be press or snap fitted into inlet 622 of second pipette tip 620, as shown in Figure 6B.
  • first pipette tip 610 is a 1-ml-pipette tip and second pipette tip 620 is a 30 ⁇ 1-pipette tip, wherein the inner diameter of outlet 624 of second pipette tip 620 is about 0.02 inches.
  • the size of outlet 624 of second pipette tip 620 is designed to be fitted into, for example, filler fluid loading port 158 of droplet actuator 150, as shown in Figure 6C.
  • Figure 6C illustrates a perspective view of two-piece pipette 600 in use with droplet actuator 150.
  • first pipette tip 610 is filled with a volume of filler fluid.
  • filler fluid drips from first pipette tip 610 into second pipette tip 620, which controls the flow rate of the filler fluid into the droplet operations gap of droplet actuator 150.
  • the filling fixtures 100 and two-piece pipette 600 are formed, for example, of plastic or glass and can be transparent, semi-transparent, or opaque.
  • the filling fixtures 100 and two-piece pipette 600 are formed, for example, by machining, injection molding, or casting. Examples of materials suitable for forming by injection molding include, but are not limited to, Delrin® 500 (available from DuPont, Wilmington, DE), polymethyl methacrylate (PMMA), polycarbonate (PC), polypropylene (PP), and polystyrene (PS).
  • Droplet actuators can include arrangements of multiple on-actuator fluid reservoirs.
  • arrangements of multiple on-actuator fluid reservoirs when fluid is dispensed from one fluid reservoir there is a risk of fluid from adjacent fluid reservoirs flooding the droplet operations gap of the droplet actuator.
  • a fluid reservoir dispensing protocol is described hereinbelow with reference to Figure 7.
  • FIG. 7 illustrates a top view of an example of an electrode arrangement 700 and a fluid reservoir dispensing protocol for improved control.
  • electrode arrangement 700 is an electrode arrangement that corresponds to an array of on-actuator fluid reservoirs (not shown) on a droplet actuator (not shown).
  • a set of three reservoir electrodes 710 is associated with each of the on-actuator fluid reservoirs of the droplet actuator.
  • each set of reservoir electrodes 710 includes reservoir electrodes 71 OA, 710B, and 710C.
  • electrode arrangement 700 includes 48 sets of reservoir electrodes 71 OA, 710B, and 7 IOC that are arranged in a 4 x 12 array.
  • the 48 sets of reservoir electrodes 71 OA, 710B, and 7 IOC are arranged in an array that includes four columns C0-C3 and twelve rows RO-Pvl l . Further, the array of reservoir electrodes 710 are fluidly connected by an arrangement of droplet operations electrodes 712 (e.g., electrowetting electrodes), as shown in Figure 7.
  • droplet operations electrodes 712 e.g., electrowetting electrodes
  • embodiments of the invention include a fluid reservoir dispensing protocol that reduces, preferably entirely eliminates, this flooding problem.
  • the reservoir electrode 710B of each of the other sets of reservoir electrodes 71 OA, 71 OB, and 7 IOC in the same row is activated. In so doing, the liquid is retained at the non-dispensing sets of reservoir electrodes 71 OA, 71 OB, and 7 IOC.
  • the reservoir electrode 71 OB at each of the locations R3-C1, R3-C2, and R3-C3 is activated.
  • liquid is retained atop the reservoir electrodes 710A, 710B, and 710C at locations R3-C1, R3-C2, and R3-C3, while liquid atop reservoir electrodes 71 OA, 71 OB, and 7 IOC at location R3-C0 is dispensed onto droplet operations electrodes 712 using a certain droplet operations sequence.
  • FIG. 8 illustrates a functional block diagram of an example of a microfluidics system 800 that includes a droplet actuator 805.
  • Digital micro fluidic technology conducts droplet operations on discrete droplets in a droplet actuator, such as droplet actuator 805, by electrical control of their surface tension (electro wetting).
  • the droplets may be sandwiched between two substrates of droplet actuator 805, a bottom substrate and a top substrate separated by a droplet operations gap.
  • the bottom substrate may include an arrangement of electrically addressable electrodes.
  • the top substrate may include a reference electrode plane made, for example, from conductive ink or indium tin oxide (ITO).
  • ITO indium tin oxide
  • the bottom substrate and the top substrate may be coated with a hydrophobic material. Droplet operations are conducted in the droplet operations gap.
  • the space around the droplets may be filled with an immiscible inert fluid, such as silicone oil, to prevent evaporation of the droplets and to facilitate their transport within the device.
  • an immiscible inert fluid such as silicone oil
  • Other droplet operations may be effected by varying the patterns of voltage activation; examples include merging, splitting, mixing, and dispensing of droplets.
  • Droplet actuator 805 may be designed to fit onto an instrument deck (not shown) of microfluidics system 800.
  • the instrument deck may hold droplet actuator 805 and house other droplet actuator features, such as, but not limited to, one or more magnets and one or more heating devices.
  • the instrument deck may house one or more magnets 810, which may be permanent magnets.
  • the instrument deck may house one or more electromagnets 815. Magnets 810 and/or electromagnets 815 are positioned in relation to droplet actuator 805 for immobilization of magnetically responsive beads.
  • the positions of magnets 810 and/or electromagnets 815 may be controlled by a motor 820.
  • the instrument deck may house one or more heating devices 825 for controlling the temperature within, for example, certain reaction and/or washing zones of droplet actuator 805.
  • heating devices 825 may be heater bars that are positioned in relation to droplet actuator 805 for providing thermal control thereof.
  • a controller 830 of microfluidics system 800 is electrically coupled to various hardware components of the invention, such as droplet actuator 805, electromagnets 815, motor 820, and heating devices 825, as well as to a detector 835, an impedance sensing system 840, and any other input and/or output devices (not shown). Controller 830 controls the overall operation of microfluidics system 800. Controller 830 may, for example, be a general purpose computer, special purpose computer, personal computer, or other programmable data processing apparatus. Controller 830 serves to provide processing capabilities, such as storing, interpreting, and/or executing software instructions, as well as controlling the overall operation of the system. Controller 830 may be configured and programmed to control data and/or power aspects of these devices. For example, in one aspect, with respect to droplet actuator 805, controller 830 controls droplet manipulation by activating/deactivating electrodes.
  • detector 835 may be an imaging system that is positioned in relation to droplet actuator 805.
  • the imaging system may include one or more light-emitting diodes (LEDs) (i.e., an illumination source) and a digital image capture device, such as a charge-coupled device (CCD) camera.
  • LEDs light-emitting diodes
  • CCD charge-coupled device
  • Impedance sensing system 840 may be any circuitry for detecting impedance at a specific electrode of droplet actuator 805.
  • impedance sensing system 840 may be an impedance spectrometer.
  • Impedance sensing system 840 may be used to monitor the capacitive loading of any electrode, such as any droplet operations electrode, with or without a droplet thereon.
  • suitable capacitance detection techniques see Sturmer et al., U.S. Patent Application Publication No. US20100194408, entitled “Capacitance Detection in a Droplet Actuator," published on Aug. 5, 2010; and Bourn et al., U.S. Patent Publication No.
  • Droplet actuator 805 may include disruption device 845.
  • Disruption device 845 may include any device that promotes disruption (lysis) of materials, such as tissues, cells and spores in a droplet actuator.
  • Disruption device 845 may, for example, be a sonication mechanism, a heating mechanism, a mechanical shearing mechanism, a bead beating mechanism, physical features incorporated into the droplet actuator 805, an electric field generating mechanism, a thermal cycling mechanism, and any combinations thereof.
  • Disruption device 845 may be controlled by controller 830.
  • aspects of the invention may be embodied as a method, system, computer readable medium, and/or computer program product.
  • aspects of the invention may take the form of hardware embodiments, software embodiments (including firmware, resident software, micro-code, etc.), or embodiments combining software and hardware aspects that may all generally be referred to herein as a "circuit,” “module” or “system.”
  • the methods of the invention may take the form of a computer program product on a computer-usable storage medium having computer-usable program code embodied in the medium.
  • the computer-usable or computer-readable medium may be, for example but not limited to, an electronic, magnetic, optical, electromagnetic, infrared, or semiconductor system, apparatus, device, or propagation medium.
  • the computer readable medium may include transitory and/or non-transitory embodiments.
  • the computer- readable medium would include some or all of the following: an electrical connection having one or more wires, a portable computer diskette, a hard disk, a random access memory (RAM), a read-only memory (ROM), an erasable programmable read-only memory (EPROM or Flash memory), an optical fiber, a portable compact disc read-only memory (CD-ROM), an optical storage device, a transmission medium such as those supporting the Internet or an intranet, or a magnetic storage device.
  • RAM random access memory
  • ROM read-only memory
  • EPROM or Flash memory erasable programmable read-only memory
  • CD-ROM compact disc read-only memory
  • CD-ROM compact disc read-only memory
  • a transmission medium such as those supporting the Internet or an intranet, or a magnetic storage device.
  • the computer-usable or computer-readable medium could even be paper or another suitable medium upon which the program is printed, as the program can be electronically captured, via, for instance, optical scanning of the paper or other medium, then compiled, interpreted, or otherwise processed in a suitable manner, if necessary, and then stored in a computer memory.
  • a computer-usable or computer-readable medium may be any medium that can contain, store, communicate, propagate, or transport the program for use by or in connection with the instruction execution system, apparatus, or device.
  • Program code for carrying out operations of the invention may be written in an object oriented programming language such as Java, Smalltalk, C++ or the like.
  • the program code for carrying out operations of the invention may also be written in conventional procedural programming languages, such as the "C" programming language or similar programming languages.
  • the program code may be executed by a processor, application specific integrated circuit (ASIC), or other component that executes the program code.
  • the program code may be simply referred to as a software application that is stored in memory (such as the computer readable medium discussed above).
  • the program code may cause the processor (or any processor-controlled device) to produce a graphical user interface ("GUI").
  • GUI graphical user interface
  • the graphical user interface may be visually produced on a display device, yet the graphical user interface may also have audible features.
  • the program code may operate in any processor-controlled device, such as a computer, server, personal digital assistant, phone, television, or any processor- controlled device utilizing the processor and/or a digital signal processor.
  • the program code may locally and/or remotely execute.
  • the program code for example, may be entirely or partially stored in local memory of the processor-controlled device.
  • the program code may also be at least partially remotely stored, accessed, and downloaded to the processor-controlled device.
  • a user's computer for example, may entirely execute the program code or only partly execute the program code.
  • the program code may be a stand-alone software package that is at least partly on the user's computer and/or partly executed on a remote computer or entirely on a remote computer or server. In the latter scenario, the remote computer may be connected to the user's computer through a communications network.
  • the invention may be applied regardless of networking environment.
  • the communications network may be a cable network operating in the radio- frequency domain and/or the Internet Protocol (IP) domain.
  • IP Internet Protocol
  • the communications network may also include a distributed computing network, such as the Internet (sometimes alternatively known as the "World Wide Web"), an intranet, a local-area network (LAN), and/or a wide-area network (WAN).
  • the communications network may include coaxial cables, copper wires, fiber optic lines, and/or hybrid-coaxial lines.
  • the communications network may even include wireless portions utilizing any portion of the electromagnetic spectrum and any signaling standard (such as the IEEE 802 family of standards, GSM/CDMA/TDMA or any cellular standard, and/or the ISM band).
  • the communications network may even include powerline portions, in which signals are communicated via electrical wiring.
  • the invention may be applied to any wireless/wireline communications network, regardless of physical componentry, physical configuration, or communications standard(s).
  • the program code may also be stored in a computer-readable memory that can direct the processor, computer, or other programmable data processing apparatus to function in a particular manner, such that the program code stored in the computer-readable memory produce or transform an article of manufacture including instruction means which implement various aspects of the method steps.
  • the program code may also be loaded onto a computer or other programmable data processing apparatus to cause a series of operational steps to be performed to produce a processor/computer implemented process such that the program code provides steps for implementing various functions/acts specified in the methods of the invention.

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  • Health & Medical Sciences (AREA)
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  • General Health & Medical Sciences (AREA)
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Abstract

L'invention concerne des mécanismes et des procédés de chargement d'un actionneur de gouttelettes avec un fluide de remplissage. Ces mécanismes et procédés sont utiles pour charger un fluide de remplissage dans un actionneur de gouttelettes d'une manière suffisamment rapide pour une utilisation pratique mais suffisamment lente pour réduire, voire empêcher le piégeage d'air dans jour d'opérations de gouttelettes pendant le processus de chargement.
PCT/US2013/068080 2012-11-02 2013-11-01 Mécanismes et procédés de chargement d'un actionneur de gouttelettes avec un fluide de remplissage WO2014078100A1 (fr)

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US10059922B2 (en) 2014-07-31 2018-08-28 Becton, Dickinson And Company Methods and systems for separating components of a biological sample with gravity sedimentation
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US10695762B2 (en) 2015-06-05 2020-06-30 Miroculus Inc. Evaporation management in digital microfluidic devices
US10464067B2 (en) 2015-06-05 2019-11-05 Miroculus Inc. Air-matrix digital microfluidics apparatuses and methods for limiting evaporation and surface fouling
US11471888B2 (en) 2015-06-05 2022-10-18 Miroculus Inc. Evaporation management in digital microfluidic devices
US11097276B2 (en) 2015-06-05 2021-08-24 mirOculus, Inc. Air-matrix digital microfluidics apparatuses and methods for limiting evaporation and surface fouling
US11944974B2 (en) 2015-06-05 2024-04-02 Miroculus Inc. Air-matrix digital microfluidics apparatuses and methods for limiting evaporation and surface fouling
US11890617B2 (en) 2015-06-05 2024-02-06 Miroculus Inc. Evaporation management in digital microfluidic devices
US10596572B2 (en) 2016-08-22 2020-03-24 Miroculus Inc. Feedback system for parallel droplet control in a digital microfluidic device
US11298700B2 (en) 2016-08-22 2022-04-12 Miroculus Inc. Feedback system for parallel droplet control in a digital microfluidic device
CN107966808A (zh) * 2016-10-19 2018-04-27 夏普生命科学（欧洲）有限公司 将流体装载至微流体设备中
EP3311918A1 (fr) * 2016-10-19 2018-04-25 Sharp Life Science (EU) Limited Chargement de fluide dans un dispositif microfluidique
US10596568B2 (en) 2016-10-19 2020-03-24 Sharp Life Science (Eu) Limited Fluid loading into a microfluidic device
US20180104687A1 (en) * 2016-10-19 2018-04-19 Sharp Life Science (Eu) Limited Fluid loading into a microfluidic device
US11253860B2 (en) 2016-12-28 2022-02-22 Miroculus Inc. Digital microfluidic devices and methods
US11833516B2 (en) 2016-12-28 2023-12-05 Miroculus Inc. Digital microfluidic devices and methods
US11623219B2 (en) 2017-04-04 2023-04-11 Miroculus Inc. Digital microfluidics apparatuses and methods for manipulating and processing encapsulated droplets
US11413617B2 (en) 2017-07-24 2022-08-16 Miroculus Inc. Digital microfluidics systems and methods with integrated plasma collection device
US11857969B2 (en) 2017-07-24 2024-01-02 Miroculus Inc. Digital microfluidics systems and methods with integrated plasma collection device
US11311882B2 (en) 2017-09-01 2022-04-26 Miroculus Inc. Digital microfluidics devices and methods of using them
US11992842B2 (en) 2018-05-23 2024-05-28 Miroculus Inc. Control of evaporation in digital microfluidics
CN113661005A (zh) * 2018-11-27 2021-11-16 斯蒂拉科技公司 微流控芯片中优化样品加载的孔
US11738345B2 (en) 2019-04-08 2023-08-29 Miroculus Inc. Multi-cartridge digital microfluidics apparatuses and methods of use
US11524298B2 (en) 2019-07-25 2022-12-13 Miroculus Inc. Digital microfluidics devices and methods of use thereof
US11772093B2 (en) 2022-01-12 2023-10-03 Miroculus Inc. Methods of mechanical microfluidic manipulation
US11857961B2 (en) 2022-01-12 2024-01-02 Miroculus Inc. Sequencing by synthesis using mechanical compression

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