Bio-Rad Laboratories, Inc.

Han, Chunxiao

Hiddessen, Amy, L.

Hindson, Benjamin, J.

Chia, Erin, R.

Lowe, Adam

Amy L Hiddessen

Benjamin J Hindson

Erin Rae Chia

Kevin D Ness

Donald Masquelier

Anthony Makarewicz

Chunxiao Han

Adam Lowe

Christopher Hindson

Geoff McDermott

Yaqi Wang

Ben Hindson

Anthony J Makarewicz

Erin R Steenblock

Donald A Masquelier

Chris Hindson

Erin Chia

System, including methods, apparatus, compositions, and kits, for making and using a stabilized emulsion. A method of generating a stabilized emulsion is provided. In the method, an aqueous phase may be provided. The aqueous phase may include an effective concentration of one or more skin-forming proteins. An emulsion may be formed. The emulsion may include droplets of a dispersed phase disposed in a continuous phase, with the aqueous phase being the continuous phase or the dispersed phase. The emulsion may be heated to create an interfacial skin between each droplet and the continuous phase, to transform the droplets into capsules.

Emulsion chemistry for encapsulated droplets

Methods, devices, systems and compositions for detecting nucleic acids in polymerase chain reaction assays, such as droplet digital polymerase chain reaction (ddPCR) assays, using intercalating dyes. A dual surfactant system with at least one fluorosurfactant and at least one non-ionic non-fluorosurfactant may be employed for droplet generation and nucleic acid detection.

Compositions, methods and systems for polymerase chain reaction assays

Method of detecting a signal from droplets for instrument calibration. In the method, a calibration standard for a droplet detection instrument may be received. The calibration standard may have been shipped at least one kilometer when received. The calibration standard may include a mixture of first droplets and second droplets. Each of the first and second droplets may be encapsulated by an immiscible carrier liquid and may have a stabilizing droplet skin that is formed by heating and that includes a skin-forming protein. Each the first droplets may contain a fluorescent dye, and each of the second droplets may contain a fluorescent dye. A fluorescence signal may be detected from a plurality of the first and second droplets with a droplet detection instrument. The fluorescence signal may be stronger for the first droplets than the second droplets.

Stabilized droplets for calibration and testing

System, including methods and compositions, for making and using emulsions that include a silicone oil and a silicone surfactant. The emulsions may include aqueous droplets disposed in a continuous phase that includes a silicone oil and a silicone surfactant. The aqueous droplets may contain an analyte, optionally at partial occupancy, and/or a luminescent (e.g., photoluminescent) reporter. An assay of the analyte may be performed with the droplets. In some cases, signals may be detected from the droplets, and a characteristic of the analyte, such as an analyte level or activity, may be determined based on the signals.

Silicone surfactants for emulsion assays

A droplet detection system comprises a first channel in fluid communication with a carrier fluid reservoir and a second channel in fluid communication with a sample reservoir. The first channel and second channel can meet at an intersection. The sample reservoir can include a sample or partition thereof. During use, an emulsion comprising one or more droplets can be generated at the intersection. The emulsion flows from the intersection along a detection channel to a collection reservoir. A detection assembly that is coupled to at least a portion of the detection channel is configured to detect a signal from the one or more droplets. An energy providing member can be in thermal communication with at least one of the carrier fluid reservoir, the sample reservoir, the intersection, the detection channel and the detection assembly. The energy providing member is configured to transfer energy to the emulsion.

Devices, systems and methods for thermal control of droplet detection

The present disclosure provides methods, devices, systems and compositions for detecting nucleic acids in polymerase chain reaction assays, such as droplet digital polymerase chain reaction (ddPCR) assays. The present disclosure provides methods, devices, systems and compositions for detecting nucleic acids in ddPCR assays using intercalating dyes. A dual surfactant system with at least one fluorosurfactant and at least one non-ionic non-fluorosurfactant may be employed for droplet generation and nucleic acid detection.

System, including methods, apparatus, and kits, for forming emulsions. In an exemplary method of generating droplets, a device may be selected that includes a plurality of emulsion-formation units each including a sample well, a continuous-phase well, a droplet well, and a channel network that fluidically interconnects the wells and creates a droplet-generation region. A discrete volume of sample-containing fluid may be placed into the sample well of each emulsion-formation unit, and a discrete volume of continuous-phase fluid into the continuous-phase well of each emulsion-formation unit. Pressure may be applied to the device with a fluidics assembly after the step of placing, such that the plurality of emulsion-formation units generate droplets in parallel with one another. A pressure signal may be detected from the fluidics assembly. Application of the pressure may be stopped when the pressure signal indicates that a sample well is empty.

System for generating droplets with pressure monitoring

System, including methods, apparatus, and kits, for forming and concentrating emulsions. An exemplary system may comprise a device including a sample well configured to receive sample-containing fluid, a continuous-phase well configured to receive continuous-phase fluid, a droplet well, and a channel network interconnecting the wells. The system also may comprise an instrument configured to operatively receive the device and to create (i) a first pressure differential to produce an emulsion collected in the droplet well and (ii) a second pressure differential to decrease a volume fraction of continuous-phase fluid in the emulsion, after the emulsion has been collected in the droplet well, by selectively driving continuous-phase fluid, relative to sample-containing droplets, from the droplet well.

System for generating droplets with push-back to remove oil

System, including methods, apparatus, and kits, for forming emulsions. An exemplary system may comprise a device including a sample well configured to receive sample-containing fluid, a continuous-phase well configured to receive continuous-phase fluid, and a droplet well. The device also may include a channel network having a first channel, a second channel, and a third channel that meet one another in a droplet-generation region. The system also may comprise a holder for the device. The system further may comprise an instrument configured to operatively receive an assembly including the device and the holder and to drive sample-containing fluid from the sample well to the droplet-generation region via the first channel, continuous-phase fluid from the continuous-phase well to the droplet-generation region via the second channel, and sample-containing droplets from the droplet-generation region to the droplet well via the third channel.

System for generating droplets&#x2014;instruments and cassette

Provided herein, are droplet mixture compositions and systems and methods for forming mixtures of droplets. The system may comprise two or more droplet generation units. Each unit may include at least one first input well, a second input well, and an output well connected to the first and second input wells by channels that form a droplet generator. The combined droplet populations can be mixed, heated, and collected for multiple uses, such as for use as calibration standards for instrument testing and analysis.

Method of detection for droplet-based assays. In an exemplary method, an open end of a channel network may be placed into an emulsion. Droplets of the emulsion may be driven along a flow path from the open end, through a confluence region where a dilution fluid is introduced into the flow path from at least one dilution inlet channel to increase an average distance between droplets, and through an examination region disposed downstream of the confluence region. Light may be detected from the examination region as droplets pass through. The channel network may include a droplet inlet channel that meets the at least one dilution inlet channel at the confluence region. The droplet inlet channel may form a tapered region that is sized such that droplets leave the tapered region in single file.

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Dublin, CA, United States of America

Amy L Hiddessen