Abstract: In biosciences and related fields, it can be useful to modify surfaces of apparatuses, devices, and materials that contact biomaterials such as biomolecules and biological micro-objects. Described herein are surface modifying and surface functionalizing reagents, preparation thereof, and methods for modifying surfaces to activate T Lymphocytes.

Abstract: Systems and methods are described herein for improved droplet generation within microfluidic apparatuses. Electrowetting forces of varying configurations may be used to separate droplets from a fluidic reservoir in a reproducible and rapid manner. In many embodiments, separation of droplets from the fluidic reservoir is performed without the use of highly specialized surfactants.

Abstract: Functional assays using reporter cell assays are described which probe the activity of at least one cell of interest. The ability to probe at least one cell is provided by using the microfluidic methods, devices and kits described herein. Assays combining both reporter cell signaling as well as binding assay signaling for at least one cell is also described herein.

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 method of preparing an antibody therapeutic is provided comprising: (a) providing a dissociated cell sample from at least one solid tumor sample obtained from a patient; (b) loading the dissociated cell sample into a microfluidic device having a flow region and at least one isolation region fluidically connected to the flow region; (c) moving at least one B cell from the dissociated cell sample into at least one isolation region in the microfluidic device, thereby obtaining at least one isolated B cell; and (d) using the microfluidic device to identify at least one B cell that produces antibodies capable of binding to cancer cells. The cancer cells can be the patient's own cancer cells. Also provided are methods of treating patients, methods of labeling or detecting cancer, engineered T or NK cells comprising antibodies or fragments thereof, and engineered antibody constructs.

Abstract: In situ-generated microfluidic capture structures incorporating a solidified polymer network, methods of preparation and use, compositions and kits therefor are described. Microfluidic capture structures may be advantageously used for assays performed within the microfluidic environment, providing flexibility in assaying micro-objects such as biological cells. Assay reagents and analytes may be incorporated within the microfluidic capture structures.

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: Optically-actuated microfluidic devices permit the use of spatially-modulated light to manipulate micro-objects such as biological cells. Systems and methods are described for providing sequences of light patterns to move and direct a plurality of micro-objects within the environment of a microfluidic device. The sequenced light patterns provide improved efficiency in directing the transport of the plurality of micro-objects. Other embodiments are described.

Abstract: Microfluidic devices having an electrowetting configuration and an optimized droplet actuation surface are provided for processing biological cells, e.g., for use in nucleic acid library preparation and/or synthesis (including amplification). The devices include a dielectric layer, a hydrophobic layer covalently bonded to the dielectric layer, and a first electrode. Methods of nucleic acid library preparation and/or synthesis can involve providing reagents to cells or nucleic acids by merging appropriate droplets on a droplet actuation surface within a water-immiscible organic liquid and can be performed in the presence of appropriate surfactants. The hydrophobic layer features self-associating molecules covalently bonded to a surface of the dielectric layer in a manner that produces a densely-packed monolayer that resists intercalation and or penetration by polar molecules or species.

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: Proto-antigen-presenting surfaces and related kits, methods, and uses are provided. The proto-antigen-presenting surface can comprise a plurality of primary activating molecular ligands comprising a major histocompatibility complex (MHC) molecule configured to bind to a T cell receptor (TCR) of a T cell and a plurality of of co-activating molecular ligands each including a TCR co-activating molecule or an adjunct TCR activating molecule, wherein an exchange factor is bound to the MHC molecules. Exchange factors include, e.g., dipeptides such as GL, GF, GR, etc. Proto-antigen-presenting surfaces can be used to rapidly prepare antigen-presenting surfaces comprising one or more peptide antigens of interest by contacting the proto-antigen-presenting surface with one or more peptide antigens so as to displace the exchange factor. As such, the disclosure facilitates rapid evaluation of the immunogenicity of peptide antigens for activating T lymphocytes.

Abstract: Methods of expanding T lymphocytes in a microfluidic device are provided. The methods can include introducing one or more T lymphocytes into a microfluidic device; contacting the one or more T lymphocytes with an activating agent; and perfusing culture medium through the microfluidic device for a period of time sufficient to allow the one or more T lymphocytes to undergo at least one round of mitotic cell division. The expansion can be non-specific or antigen-specific. T lymphocytes produced according to the disclosed methods are also provided, along with methods of treating cancer in a subject. The methods of treating cancer can include isolating T lymphocytes from a tissue sample obtained from the subject; expanding the isolated T lymphocytes in a microfluidic device; exporting the expanded T lymphocytes from the microfluidic device; and reintroducing the expanded T lymphocytes into the subject.

Abstract: Methods are provided for the automated detection of assay-positive assay areas in a microfluidic device. When assays are performed in a microfluidic device, the configuration of the microfluidic circuit and its constituent circuit elements can determine where the reagents/analytes used in the assay can be located within the microfluidic circuit. Methods are provided for automatic identification of the size and shape of the assay areas based on a number of parameters which may include type of assay involved, shape and dimensions of microfluidic circuit elements, velocity and physical characteristics of the fluidic medium within the microfluidic circuit, physical/chemical properties of the analytes/reagents, and/or the number of cells being assayed.

Abstract: Single-sided optoelectrowetting (SSOEW)-configured substrates are provided, as well as microfluidic devices that include such substrates. The substrates can include a planar electrode, a photoconductive (or photosensitive) layer, a dielectric layer (single-layer or composite), a mesh electrode, and a hydrophobic coating. Fluid droplets can be moved across the hydrophobic coating of such substrates in a light-actuated manner, upon the application of a suitable AC voltage potential across the substrate and the focusing of light into the photoconductive layer of the substrate in a location proximal to the droplets. Walls can be disposed upon the substrates to form the microfluidic devices. Together the walls and substrate can form a microfluidic circuit, through which droplets can be moved.

Abstract: Methods are described herein for isolating clonal populations of T 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 T cells (or precursors thereof) that have undergone a genomic editing process in corresponding sequestration pens of a microfluidic device; expanding the T cells into respective clonal populations of T cells; detecting, in one or more T cells of each clonal population, the absence of a cell surface marker that was present in the individual T cells (or precursors thereof); and detecting, in one or more T 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 T cells. Also described are compositions comprising one or more clonal populations of T cells isolated according to the methods disclosed herein.

Abstract: In biosciences and related fields, it can be useful to modify surfaces of apparatuses, devices, and materials that contact biomaterials such as biomolecules and biological micro-objects. Described herein are surface modifying and surface functionalizing reagents, preparation thereof, and methods for modifying surfaces to provide improved or altered performance with biomaterials.

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: 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 subsystem, and which can be configured to supply light of substantially uniform intensity to the light modulating subsystem or directly to the optical train.

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.