Abstract: Individual biological micro-objects can be deterministically selected and moved into holding pens in a micro-fluidic device. A flow of a first liquid medium can be provided to the pens. Physical pens can be structured to impede a direct flow of the first medium into a second medium in the pens while allowing diffusive mixing of the first medium and the second medium. Virtual pens can allow a common flow of medium to multiple ones of the pens.

Abstract: A system for operating an electrokinetic device includes a support configured to hold and operatively couple with the electrokinetic device, an integrated electrical signal generation subsystem configured to apply a biasing voltage across a pair of electrodes in the electrokinetic device, and a light modulating subsystem configured to emit structured light onto the electrokinetic device. The system can further include a thermally controlled flow controller, and/or be configured to measure impedance across the electrokinetic device. The system can be a light microscope, including an optical train. The system can further include a light pipe, which can be part of the light modulating system, and which can be configured to supply light of substantially uniform intensity to the light modulating system or directly to the optical train.

Abstract: A microfluidic device can comprise at least one swept region that is fluidically connected to unswept regions. The fluidic connections between the swept region and the unswept regions can enable diffusion but substantially no flow of media between the swept region and the unswept regions. The capability of biological micro-objects to produce an analyte of interest can be assayed in such a microfluidic device. Biological micro-objects in sample material loaded into a microfluidic device can be selected for particular characteristics and disposed into unswept regions. The sample material can then be flowed out of the swept region and an assay material flowed into the swept region. Flows of medium in the swept region do not substantially affect the biological micro-objects in the unswept regions, but any analyte of interest produced by a biological micro-object can diffuse from an unswept region into the swept region, where the analyte can react with the assay material to produce a localized detectable reaction.

Abstract: Apparatuses and methods are described for the use of optically driven bubble, convective and displacing fluidic flow to provide motive force in microfluidic devices. Alternative motive modalities are useful to selectively dislodge and displace micro-objects, including biological cells, from a variety of locations within the enclosure of a microfluidic device.

Abstract: Incubators including an enclosure with an internal chamber configured to support a cell culture plate comprising a plurality of wells are disclosed. The enclosure includes a plurality of openings configured to allow access to the wells. The incubators include a sealing element configured to seal the plurality of openings in the enclosure. The sealing element comprises a plurality of openings corresponding to at least a subset of the plurality of openings in the enclosure. Access to the internal chamber can be provided by aligning the plurality of openings in the sealing element with the plurality of openings in the enclosure. Methods for using the incubators are also provided.

Abstract: In situ-generated microfluidic isolation structures incorporating a solidified polymer network, methods of preparation and use, compositions and kits therefor are described. The ability to introduce in real time, a variety of isolating structures including pens and barriers offers improved methods of micro-object manipulation in microfluidic devices. The in situ-generated isolation structures may be permanently or temporarily installed.

Abstract: In situ-generated microfluidic isolation structures incorporating a solidified polymer network, methods of preparation and use, compositions and kits therefor are described. The ability to introduce in real time, a variety of isolating structures including pens and barriers offers improved methods of micro-object manipulation in microfluidic devices. The in situ-generated isolation structures may be permanently or temporarily installed.

Abstract: A microfluidic device can comprise at least one swept region that is fluidically connected to unswept regions. The fluidic connections between the swept region and the unswept regions can enable diffusion but substantially no flow of media between the swept region and the unswept regions. The capability of biological micro-objects to produce an analyte of interest can be assayed in such a microfluidic device. Biological micro-objects in sample material loaded into a microfluidic device can be selected for particular characteristics and disposed into unswept regions. The sample material can then be flowed out of the swept region and an assay material flowed into the swept region. Flows of medium in the swept region do not substantially affect the biological micro-objects in the unswept regions, but any analyte of interest produced by a biological micro-object can diffuse from an unswept region into the swept region, where the analyte can react with the assay material to produce a localized detectable reaction.

Abstract: Biological activity in holding pens in a micro-fluidic device can be assayed by placing in the holding pens capture objects that bind a particular material of interest produced by the biological activity. The biological material of interest that binds to each capture object can then be assessed, either in the micro-fluidic device or after exporting the capture object from the micro-fluidic device. The assessment can be utilized to characterize the biological activity in each holding pen. The biological activity can be production of the biological material of interest. Thus, the biological activity can correspond to or arise from one or more biological cells. Biological cells within a holding pen can be clonal cell colonies. The biological activity of each clonal cell colony can be assayed while maintaining the clonal status of each colony.

Abstract: Systems, methods and kits are described for culturing one or more biological cells in a microfluidic device, including provision of nutrients and gaseous components configured to enhance cell growth, viability, portability, or any combination thereof. In some embodiments, culturing a single cell may produce a clonal population in the microfluidic device.

Abstract: Incubators including an enclosure with an internal chamber configured to support a cell culture plate comprising a plurality of wells are disclosed. The enclosure includes a plurality of openings configured to allow access to the wells. The incubators include a sealing element configured to seal the plurality of openings in the enclosure. The sealing element comprises a plurality of openings corresponding to at least a subset of the plurality of openings in the enclosure. Access to the internal chamber can be provided by aligning the plurality of openings in the sealing element with the plurality of openings in the enclosure. Methods for using the incubators are also provided.

Abstract: Incubators are disclosed which include an enclosure with an internal chamber configured to support a cell culture plate and provide an environment suitable for maintaining and/or culturing biological cells. The enclosure can include one or more openings configured to allow access to the cell culture plate. The incubators can further include a structure having a plurality of openings configured to be aligned with a corresponding plurality of wells in the cell culture plate, and a sealing element configured to moveably seal the plurality of openings in the structure. The sealing element can comprise a plurality of openings corresponding to at least a subset of the plurality of openings of the structure. Access to the internal chamber can be provided by aligning the plurality of openings in the sealing element with the plurality of openings in the structure. Methods for using the incubators are also provided.

Abstract: Methods, systems and kits are described herein for detecting the results of an assay. In particular, the methods, systems and devices of the present disclosure rely on a difference between the diffusion rates of a reporter molecule and an analyte of interest in order to quantify an amount of analyte in a microfluidic device. The analyte may be a secreted product of a biological micro-object.

Abstract: Systems, methods and kits are described for culturing one or more biological cells in a microfluidic device, including provision of nutrients and gaseous components configured to enhance cell growth, viability, portability, or any combination thereof. In some embodiments, culturing a single cell may produce a clonal population in the microfluidic device.

Abstract: A microfluidic apparatus is provided having one or more sequestration pens configured to isolate one or more target micro-objects by changing the orientation of the microfluidic apparatus with respect to a globally active force, such as gravity. Methods of selectively directing the movements of micro-objects in such a microfluidic apparatus using gravitational forces are also provided. The micro-objects can be biological micro-objects, such as cells, or inanimate micro-objects, such as beads.

Abstract: Biological activity in holding pens in a micro-fluidic device can be assayed by placing in the holding pens capture objects that bind a particular material of interest produced by the biological activity. The biological material of interest that binds to each capture object can then be assessed, either in the micro-fluidic device or after exporting the capture object from the micro-fluidic device. The assessment can be utilized to characterize the biological activity in each holding pen. The biological activity can be production of the biological material of interest. Thus, the biological activity can correspond to or arise from one or more biological cells. Biological cells within a holding pen can be clonal cell colonies. The biological activity of each clonal cell colony can be assayed while maintaining the clonal status of each colony.

Abstract: Methods, systems and kits are described herein for detecting the results of an assay. In particular, the methods, systems and devices of the present disclosure rely on a difference between the diffusion rates of a reporter molecule and an analyte of interest in order to quantify an amount of analyte in a microfluidic device. The analyte may be a secreted product of a biological micro-object.

Abstract: A microfluidic apparatus can comprise a dielectrophoresis (DEP) configured section for holding a first liquid medium and selectively inducing net DEP forces in the first liquid medium. The microfluidic apparatus can also comprise an electrowetting (EW) configured section for holding a second liquid medium on an electrowetting surface and selectively changing an effective wetting property of the electrowetting surface. The DEP configured section can be utilized to select and move a micro-object in the first liquid medium. The EW configured section can be utilized to pull a droplet of the first liquid medium into the second liquid medium.

Abstract: A microfluidic device can comprise a plurality of interconnected microfluidic elements. A plurality of actuators can be positioned abutting, immediately adjacent to, and/or attached to deformable surfaces of the microfluidic elements. The actuators can be selectively actuated and de-actuated to create directed flows of a fluidic medium in the microfluidic (or nanofluidic) device. Further, the actuators can be selectively actuated and de-actuated to create localized flows of a fluidic medium in the microfluidic device to move reagents and/or micro-objects in the microfluidic device.

Abstract: Methods are described herein for isolating clonal populations of cells having a defined genetic modification. The methods are performed, at least in part, in a microfluidic device comprising one or more sequestration pens. The methods include the steps of: maintaining individual cells (or precursors thereof) that have undergone a genomic editing process in corresponding sequestration pens of a microfluidic device; expanding the individual cells into respective clonal populations of cells; and detecting, in one or more cells of each clonal population, the presence of a first nucleic acid sequence that is indicative of the presence of an on-target genome edit in the clonal population of cells. Also described are methods of performing genome editing within a microfluidic device, and compositions comprising one or more clonal populations of cells generated according to the methods disclosed herein.