Abstract: The systems and methods of the invention provide a guided approach to pyrosequencing (i.e., hybrid pyrosequencing). A de novo nucleic acid sequence may compared to a library of possible results and the next nucleotide to be dispensed is selected based on the comparison of the de novo sequence and the library of possible results. In another example, at least the first nucleotide to be dispensed is selected based on a query of a database(s) of non-sequence parameters (e.g., incidence of infection, diagnostic symptoms, sample source) and subsequent dispensations determined based on a comparison of the de novo sequence and the library of possible results (e.g., candidate sequences). The systems and methods of the invention may be performed using a droplet actuator.

Abstract: A method, circuit and apparatus for detecting capacitance on a droplet actuator, inter alia, for determining the presence, partial presence or absence of a droplet at an electrode on a droplet actuator by: (a) providing a droplet actuator comprising: (i) a substrate comprising electrodes arranged on the substrate for conducting droplet operations on a surface of the substrate; (ii) a capacitance detection circuit for detecting capacitance at the droplet operations surface at one or more of the electrodes; (b) detecting capacitance at the droplet operations surface at one or more of the electrodes; and (c) determining from the capacitance the presence, partial presence or absence of a droplet at the droplet operations surface at the electrode.

Abstract: Aspects of embodiments may include methods for automated enzymatic detection of glucose-6-phosphate dehydrogenase (G6PD) activity. Aspects of embodiments may include methods for enzymatic detection of G6PD activity in droplets in oil. Aspects of embodiments may include a system including a droplet actuator. Aspects of embodiments may include a treatment method.

Abstract: The present invention relates to droplet-based particle sorting. According to one embodiment, a droplet microactuator is provided and includes: (a) a suspension of particles; and (b) electrodes arranged for conducting droplet operations using droplets comprising particles. A method of transporting a particle is also provided, wherein the method includes providing a droplet comprising the particle and transporting the droplet on a droplet microactuator.

Abstract: Methods and devices for conducting chemical or biochemical reactions that require multiple reaction temperatures are described. The methods involve moving one or more reaction droplets or reaction volumes through various reaction zones having different temperatures on a microfluidics apparatus. The devices comprise a microfluidics apparatus comprising appropriate actuators capable of moving reaction droplets or reaction volumes through the various reaction zones.

Abstract: The present invention is directed to a droplet actuator and methods of making and using the droplet actuator including one or more substrates configured to form a droplet operations gap and including a physical or chemical feature that may be provided at a predetermined locus within or exposed to the droplet operations gap and configured to retain a bubble in position within the droplet operations gap.

Abstract: A method, circuit and apparatus for detecting capacitance on a droplet actuator, inter alia, for determining the presence, partial presence or absence of a droplet at an electrode on a droplet actuator by: (a) providing a droplet actuator comprising: (i) a substrate comprising electrodes arranged on the substrate for conducting droplet operations on a surface of the substrate; (ii) a capacitance detection circuit for detecting capacitance at the droplet operations surface at one or more of the electrodes; (b) detecting capacitance at the droplet operations surface at one or more of the electrodes; and (c) determining from the capacitance the presence, partial presence or absence of a droplet at the droplet operations surface at the electrode.

Abstract: The invention provides a method of dispersing or circulating magnetically responsive beads within a droplet in a droplet actuator. The invention, in one embodiment, makes use of a droplet actuator with a plurality of droplet operations electrodes configured to transport the droplet, and a magnetic field present at a portion of the plurality of droplet operations electrodes. A bead-containing droplet is provided on the droplet actuator in the presence of the uniform magnetic field. Beads are circulated in the droplet during incubation by conducting droplet operations on the droplet within a uniform region of the magnate field. Other embodiments are also provided.

Abstract: The invention provides droplet actuators and droplet actuator cassettes including reagent storage capabilities, as well as methods of making and using the droplet actuators and cassettes. The invention also provides continuous flow channel elements and techniques for using electrodes to manipulate droplets in flowing streams. The invention also discloses methods of separating compounds on a droplet actuator. Various other aspects of the invention are also disclosed.

Abstract: Sample processing droplet actuators, systems and methods are provided. According to one embodiment, a stamping device including a droplet microactuator is provided and includes: (a) a first plate including a path or network of control electrodes for transporting droplets on a surface thereof; (b) a second plate mounted in a substantially parallel orientation with respect to the first plate providing an interior volume between the plates, the second plate including one or more stamping ports for transporting some portion or all of a droplet from the interior volume to an exterior location; (c) a port for introducing fluid into the interior volume between the plates; and (d) a path or network of reference electrodes corresponding to the path or network of control electrodes. Associated systems and methods including the stamping device are also provided.

Abstract: The present invention provides for the detection of cardiac markers on a droplet actuator. An aspect provides a method of assaying a cardiac marker in a biological sample from a subject, the method including providing a droplet actuator, loading the biological sample and assay reagents on the droplet actuator, executing droplet operations to create sample droplets from the sample and reagent droplets from the reagents on the droplet actuator, and executing droplet operations using the sample droplets and reagent droplets to produce a detectable signal indicative of the quantity of the cardiac marker in the biological sample. Still other aspects are provided.

Abstract: The present invention relates to bead incubating and washing on a droplet actuator. Methods for incubating magnetically responsive beads that are labeled with primary antibody, a sample (i.e., analyte), and secondary reporter antibodies on a magnet, on and off a magnet, and completely off a magnet are provided. Also provided are methods for washing magnetically responsive beads using shape-assisted merging of droplets. Also provided are methods for shape-mediated splitting, transporting, and dispensing of a sample droplet that contains magnetically responsive beads. The apparatuses and methods of the invention provide for rapid time to result and optimum detection of an analyte in an immunoassay.

Abstract: A series of microactuators for manipulating small quantities of liquids, and methods of using these for manipulating liquids, are disclosed. The microactuators are based on the phenomenon of electrowetting and contain no moving parts. The force acting on the liquid is a potential-dependent gradient of adhesion energy between the liquid and a solid insulating surface.

Abstract: A method is provided for conducting a droplet-based enzymatic assay, e.g., for diagnostic purposes. On a droplet actuator, a droplet comprising an enzyme of interest is provided along with a droplet comprising a substrate which is potentially modified in the presence of the enzyme. Droplet operations are executed to combine the enzyme and substrate droplets on the droplet actuator, thereby yielding an assay droplet on the droplet actuator. Detecting modification of the substrate by the enzyme in the assay droplet occurs on the droplet actuator. Modified substrate preparations for conducting such enzymatic assays are also provided.

Abstract: Droplet actuators and techniques for droplet-based assays are provided. A method of conducting an assay comprises: incubating a droplet in oil, the droplet comprising an umbelliferone substrate, a sample potentially comprising an enzyme which cleaves the umbelliferone substrate, and a zwitterionic surfactant; and detecting a signal emitted from the droplet.

Abstract: The present invention relates to filler fluids for droplet operations. According to one embodiment of this aspect, a droplet microactuator including an opaque filler fluid and a transparent droplet thereon is provided. The droplet microactuator may further include a first substrate comprising electrodes configured to effect electrowetting mediated droplet operations on a surface of the first substrate; a second substrate spaced apart from the surface of the first substrate by a distance sufficient to define an interior volume between the first substrate and second substrate; and wherein the droplet and the filler fluid are in the interior volume, and wherein the droplet is surrounded by the filler fluid and arranged with respect to the electrodes in a manner which permits electrowetting mediated droplet operations to be effected on the droplet using the electrodes.

Abstract: Droplet actuator apparatus and system are provided. An apparatus comprises: a microfluidics system having an insertion slot for insertion of a droplet actuator; a bottom plate against which the droplet actuator would slide when inserting the droplet actuator into the insertion slot; and means for forcing a substrate in the droplet actuator parallel to the bottom plate, wherein warpage in the substrate is corrected.

Abstract: The invention provides a method of dispersing or circulating magnetically responsive beads within a droplet in a droplet actuator. The invention, in one embodiment, makes use of a droplet actuator with a plurality of droplet operations electrodes configured to transport the droplet, and a magnetic field present at a portion of the plurality of droplet operations electrodes. A bead-containing droplet is provided on the droplet actuator in the presence of the uniform magnetic field. Beads are circulated in the droplet during incubation by conducting droplet operations on the droplet within a uniform region of the magnetic field wherein droplet operations do not allow magnetically responsive beads to be introduced into a region of the magnetic field which is sufficiently non-uniform to cause bead clumping resulting in a more homogenous distribution of the beads in the droplet. Other embodiments are also provided.

Abstract: The present invention is directed to droplet actuators with local variation in gap height and methods of their use to facilitate droplet splitting and merging operations. The droplet actuators have increased gap-height regions that track droplet transport paths such that droplets can be transported along the paths with reduced risk of merging with droplets on adjacent paths.

Abstract: The invention provides a droplet actuator. The droplet actuator may include a base substrate and a top substrate separated to form a gap. The base substrate may include electrodes configured for conducting droplet operations in the gap; and the top substrate may include a glass substrate portion coupled to a non-glass portion, where the non-glass portion may include one or more openings establishing a fluid path extending from an exterior of the droplet actuator and into the gap. The invention also provides related methods of manufacturing the droplet actuator, methods of using the droplet actuator, and methods of loading the droplet actuator.