Devices and methods for analyzing biological samples

Abstract

Systems and methods for analyzing biological samples are described herein.A method for processing an analyte includes the steps of providing a fluidic device containing the analyte and one or more polymer precursors, selecting a distinct area within the fluidic device, providing an energy source in optical communication with the fluidic device, and selectively providing units of energy generated from the energy source to the fluidic device to generate a polymer matrix within the fluidic device, wherein the polymer matrix is ​​within or adjacent to the distinct area.

Description

[Technical Field]

[0001] cross reference This application claims the benefit of U.S. Provisional Application No. 63 / 135,463, filed on 8 January 2021, which is incorporated herein by reference in its entirety.

[0002] Embedding by reference All publications, patents, and patent applications referenced herein are incorporated by reference to the same extent that each individual publication, patent, or patent application is specifically and individually indicated as being incorporated by reference. To the extent that any publications, patents, or patent applications incorporated by reference conflict with any disclosures contained herein, this specification is intended to supersede and / or take precedence over any such conflicting material. [Background technology]

[0003] background In the field of cell biology, single-cell analysis can include the study of genomics, transcriptomics, proteomics, metabolomics, and intercellular interactions at the single-cell level. Due to the heterogeneity found in both eukaryotic and prokaryotic cell populations, analyzing single cells can make it possible to discover mechanisms not seen when studying bulk populations of cells. Single-cell analysis allows for tracking or observing single-cell changes at the level of genes, proteins, or other cellular components. For example, in cancers where cells may mutate, the goal may be to understand how the cancer is changing at the genetic level. These patterns of somatic mutations and copy number abnormalities can be observed using single-cell sequencing. [Overview of the project] [Means for solving the problem]

[0004] overview Here, the need to compartmentalize the components of a biological sample is recognized in order to perform one or more assays on individual components within the compartment. One or more assays can be performed with or without the need for additional processing of individual components (e.g., without the need for a nucleotide amplification step) while preserving the spatial information of the individual components. Compartments can be generated or dismantled on demand to localize (i.e., confine to) or release targeted components of the biological sample.

[0005] This specification provides a method comprising the steps of: providing a fluid device containing an analyte and one or more polymer precursors; identifying a distinct area within the fluid device; and selectively supplying units of energy generated from an energy source to the fluid device to generate a polymer matrix from the one or more polymer precursors within the fluid device, wherein the polymer matrix is ​​located within or adjacent to the distinct area. In some embodiments, the fluid device includes a flow channel. In some embodiments, the fluid device includes an open configuration, e.g., the open configuration shown in the embodiment of Figure 20. In some embodiments, the fluid device contains one or more distinct locations, where one or more distinct locations are not in fluid communication with another distinct location. In some embodiments, one or more distinct locations are one or more wells on the surface of a plate. In some embodiments, where the distinct locations are bounded by wells or cavities or containers in or on the surface, one or more distinct locations are open at the top. That is, in some embodiments, one or more distinct locations contain an analyte or biological component. In some embodiments, the units of energy are an amount of energy, e.g., a amount of light energy, that is effective in causing the synthesis of a chamber in the channel. The unit values ​​of energy for a given embodiment can vary widely, but are not limited, depending on factors such as the characteristics of the spatial energy modulating element, the size of the channel, the size and geometry of the desired chamber, and the characteristics of the polymer precursor.

[0006] In some embodiments, the fluid device further includes a space energy modulating element. In some embodiments, the fluid device includes a surface on which a capture probe is immobilized, the capture probe coupling to the analyte to immobilize the analyte on the surface. In some embodiments, a polymer matrix is ​​generated adjacent to or surrounding the analyte to immobilize the analyte. In some embodiments, the polymer matrix forms a hydrogel. In some embodiments, the capture probe includes one or more functional groups that can interact with the analyte. In some embodiments, one or more functional groups include a DNA sequence complementary to the target DNA or RNA.

[0007] In some embodiments, the energy source is in optical communication with the fluid device. In some embodiments, the spatial energy modulator is a photogenerating device, such as a digital micromirror device. In some embodiments, the photogenerating device generates light in the 350 nm to 800 nm range. In some embodiments, the photogenerating device generates light in the 350 nm to 600 nm range. In some embodiments, the photogenerating device generates light in the 350 nm to 450 nm range. In some embodiments, the photogenerating device generates UV light. In some embodiments, the step of selectively supplying units of energy generated from the energy source to the fluid device to generate a polymer matrix within the fluid device is performed using a spatial energy modulator, which is a spatial light modulator (SLM). In some embodiments, the SLM is a digital micromirror device (DMD). In some embodiments, the SLM is a laser beam guided using a galvanometer. In some embodiments, the SLM is liquid crystal based. Using the light energy from the spatial light modulator or spatial energy modulator, the polymer precursor may be photocrosslinked to form polymer matrix walls that create chambers formed in the channel.

[0008] In some embodiments, the area of ​​the separate area is less than the area of ​​the fluid device. In some embodiments, the analyte is captured within the separate area. In some embodiments, the size and shape of the separate area are adjustable according to the size, shape, or other properties of the analyte. In some embodiments, an algorithm is used to determine the shape and size of the separate area. In some embodiments, the algorithm is a supervised learning algorithm, a self-supervised learning algorithm, or an unsupervised learning algorithm. In some embodiments, the separate area is optically identified. In some embodiments, the separate area is identified using a detector configured to detect the location of the analyte from an image of light collected from the fluid device. In some embodiments, the detector configured to detect the location of the analyte within the fluid device is a microscope objective lens for imaging the fluid device. In some embodiments, an algorithm is used to determine where the analyte is located based on the imaging. In some embodiments, the imaging is bright-field imaging, phase-contrast imaging, or fluorescence imaging, or any combination thereof. In some embodiments, the algorithm is a supervised learning algorithm, a self-supervised learning algorithm, or an unsupervised learning algorithm. In some embodiments, the objective lens is coupled to an energy source that radiates energy to a separate area in a fluid device.

[0009] In some embodiments, the method further includes the step of introducing one or more reagents into a polymer matrix that reacts with the analyte. In some embodiments, one or more reagents flow through a membrane. In some embodiments, the membrane is semipermeable. In some embodiments, the membrane contains pores. In some embodiments, the pores are less than 10 μm. In some embodiments, one or more reagents include: one or more enzymes, drug molecules, oligonucleotides, primers, or any combination thereof. In some embodiments, one or more reagents are lysis reagents, e.g., lysis reagents for lysing cells to be selectively removed from channels of a fluid device. In some embodiments, one or more reagents are nucleic acid denaturation reagents. In some embodiments, one or more reagents degrade the polymer matrix.

[0010] In some embodiments, the analyte is a cellular component. In some embodiments, the analyte is a small molecule consisting of nucleic acids, amino acids, intracellular proteins, surface proteins, secretory proteins, exosomes, metabolites or lipids, or any combination thereof. In some embodiments, the analyte is captured by a capture probe within a polymer matrix. In some embodiments, the analyte is captured by a capture probe on the surface of a polymer matrix.

[0011] In some embodiments, the analyte is released from the cell upon interaction with the reagent. In some embodiments, the reagent is an oxidizing or reducing agent. In some embodiments, the reagent is an organic or inorganic molecule. In some embodiments, the organic or inorganic molecule is a pharmaceutical compound or a surfactant. In some embodiments, the reagent is a protein. In some embodiments, the reagent is a DNA aptamer. In some embodiments, the reagent is a bead containing a biomolecule. In some embodiments, the reagent is a biological species. In some embodiments, the biological species is a virus or a cell.

[0012] In some embodiments, the analyte is released from the cells upon exposure to an energy source. In some embodiments, the energy source is UV light for lysing the cells. In some embodiments, the energy source is visible light for lysing the cells. In some embodiments, UV light is used to activate a photoactivatable surfactant and lyse the cells. In some embodiments, visible light is used to activate a photoactivatable surfactant and lyse the cells.

[0013] In some embodiments, the method further includes the step of identifying an analyte or its components. In some embodiments, the analyte is a nucleic acid, amino acid, intracellular protein, surface protein, secretory protein, exosome, metabolite or lipid, or any combination thereof. In some embodiments, the analyte is a nucleic acid molecule, and the identification step includes sequencing of the nucleic acid molecule or its derivatives.

[0014] In some embodiments, the method further includes a step of measuring the quality of the analyte or its components. In some embodiments, quality is the shape or size of the analyte or its components. In some embodiments, the method further includes a step of performing one or more functional assays to analyze cells or their components to evaluate cell viability, cell morphology, cell secretion, cell response, cell-cell interactions, or any combination thereof. In some embodiments, one or more functional assays are colorimetric assays or fluorescence assays. In some embodiments, one or more functional assays are performed using bright-field phase imaging or fluorescence imaging of the analyte.

[0015] In some embodiments, the method further includes the step of performing one or more omics assays to characterize and quantify cells or their components. In some embodiments, one or more omics assays are proteomics assays, transcriptomics assays, genomics assays, or epigenomics assays, or any combination thereof. In some embodiments, one or more omics assays are multi-omics assays.

[0016] Another aspect of the present disclosure provides a method for processing an analyte, comprising the steps of: providing a fluid device comprising the analyte and one or more polymer precursors; and configuring a digital micromirror device such that units of energy generated from an energy source are directed to separate areas of the fluid device to generate a polymer matrix from the one or more polymer precursors within the fluid device, wherein the polymer matrix comprises or encapsulates the analyte.

[0017] In some embodiments, the fluid device includes a flow channel (which may be referred to herein as “channel”). In some embodiments, the fluid device includes an open configuration. In some embodiments, the fluid device includes one or more distinct locations, one or more of which are not in fluid communication with another distinct location. In some embodiments, one or more distinct locations are one or more well plates. In some embodiments, one or more distinct locations are open at the top. In some embodiments, one or more distinct locations contain an analyte.

[0018] In some embodiments, the separate area is adjacent to or surrounding the analyte. In some embodiments, the separate area is adjustable according to the size, shape, or other properties of the analyte. In some embodiments, the separate area is optically identified. In some embodiments, the detector is configured to detect the location of the analyte within a fluid device. In some embodiments, the detector configured to detect the location of the analyte within a fluid device is a microscope objective lens for imaging the fluid device via an image of the fluid channel. In some embodiments, an algorithm is used to determine, based on the imaging, where the analyte or biological component is located. In some embodiments, the imaging is bright-field imaging, phase-contrast imaging, or fluorescence imaging, or any combination thereof. In some embodiments, the algorithm is a supervised learning algorithm, a self-supervised learning algorithm, or an unsupervised learning algorithm. In some embodiments, the objective lens is coupled to an energy source that is optically communicating with the fluid device. In some embodiments, a digital micromirror device directs units of energy to a fluid device based on information extracted by an algorithm from an image of a fluid channel.

[0019] In some embodiments, the method further includes the step of introducing one or more reagents into a polymer matrix that reacts with the analyte. In some embodiments, one or more reagents flow through a membrane. In some embodiments, the membrane is semipermeable. In some embodiments, the membrane contains pores. In some embodiments, the pores are less than 10 μm. In some embodiments, one or more reagents are enzymes, oligonucleotides, primers, or any combination thereof. In some embodiments, one or more reagents are dissolving reagents. In some embodiments, one or more reagents are nucleic acid denaturing reagents. In some embodiments, one or more reagents degrade the polymer matrix.

[0020] In some embodiments, the analyte is a component of a cell. In some embodiments, the cell component is a small molecule composed of nucleic acids, amino acids, intracellular proteins, surface proteins, secreted proteins, exosomes, metabolites or lipids, or any combination thereof. In some embodiments, the analyte is captured by a capture probe within a polymeric matrix. In some embodiments, the analyte is captured by a capture probe on the surface of the polymeric matrix.

[0021] In some embodiments, the analyte is released from the cell upon interaction with a reagent. In some embodiments, the reagent is an oxidizing agent or a reducing agent. In some embodiments, the reagent is an organic molecule or an inorganic molecule. In some embodiments, the organic molecule or the inorganic molecule is a pharmaceutical compound or a surfactant. In some embodiments, the reagent is a protein. In some embodiments, the reagent is a DNA aptamer. In some embodiments, the reagent is a bead with a biomolecule. In some embodiments, the reagent is a biological species. In some embodiments, the biological species is a virus or a cell.

[0022] In some embodiments, the analyte is released from the cell upon exposure to an energy source. In some embodiments, the energy source is UV light for lysing the cell. In some embodiments, the energy source is visible light for lysing the cell. In some embodiments, UV light is used to activate a photoactivated surfactant to lyse the cell. In some embodiments, visible light is used to activate a photoactivated surfactant to lyse the cell.

[0023] In some embodiments, the method further comprises the step of identifying the analyte or its derivative. In some embodiments, the analyte is a nucleic acid molecule, and the identifying step comprises sequencing the nucleic acid molecule or its derivative. In some embodiments, the method further comprises the step of measuring the quality of the analyte. In some embodiments, the quality is the shape or size of the analyte.

[0024] In some embodiments, the method further includes the step of performing one or more functional assays to analyze the analytes to evaluate cell viability, cell morphology, cell secretion, cell response, cell-cell interactions, or any combination thereof. In some embodiments, one or more functional assays are colorimetric assays or fluorescence assays. In some embodiments, one or more functional assays are performed using bright-field phase imaging or fluorescence imaging of the analytes.

[0025] In some embodiments, the method further includes the step of performing one or more omics assays to characterize and quantify the analyte or its components. In some embodiments, one or more omics assays are proteomics assays, transcriptomics assays, genomics assays, or epigenomics assays, or any combination thereof. In some embodiments, one or more omics assays are multi-omics assays.

[0026] Another aspect of the present disclosure provides a method for processing an analyte, comprising the steps of: providing a fluid device comprising the analyte and one or more polymer precursors; and using one or more polymer precursors within the fluid device to generate a polymer matrix from the one or more polymer precursors, wherein the polymer matrix comprises the analyte, and the generation of the polymer matrix is ​​carried out in the absence of a physical photomask.

[0027] In some embodiments, the fluid device includes a flow channel. In some embodiments, the fluid device includes an open configuration. In some embodiments, the fluid device includes one or more distinct locations, one or more of which are not in fluid communication with another distinct location. In some embodiments, one or more distinct locations are one or more well plates. In some embodiments, one or more distinct locations are open at the top. In some embodiments, one or more distinct locations contain an analyte.

[0028] In some embodiments, the fluid device further comprises one or more monomers. In some embodiments, the fluid device further comprises a space energy modulating element. In some embodiments, the fluid device comprises a surface on which a capture probe is immobilized, the capture probe coupling to the analyte to immobilize the analyte on the surface. In some embodiments, a polymer matrix is ​​generated adjacent to or surrounding the analyte to immobilize the analyte. In some embodiments, the polymer matrix forms a hydrogel. In some embodiments, the capture probe comprises one or more functional groups that can interact with the analyte. In some embodiments, one or more functional groups comprise a DNA sequence complementary to the target DNA or RNA.

[0029] In some embodiments, the generation of a polymer matrix within the fluid device includes exposing one or more polymer precursors to an energy source. In some embodiments, the energy source is a photogenerator. In some embodiments, the photogenerator generates light in the 350 nm to 800 nm range. In some embodiments, the photogenerator generates light in the 350 nm to 600 nm range. In some embodiments, the photogenerator generates light in the 350 nm to 450 nm range. In some embodiments, the photogenerator generates UV light. In some embodiments, the generation of a polymer matrix within the fluid device is performed using a spatial light modulator (SLM). In some embodiments, the SLM is a digital micromirror device (DMD). In some embodiments, the SLM is a laser beam guided using a galvanometer. In some embodiments, the SLM is liquid crystal based.

[0030] In some embodiments, the polymer matrix is ​​generated in a separate area of ​​the fluid device. In some embodiments, the separate area is adjacent to or surrounds the analyte. In some embodiments, the area of ​​the separate area is less than the area of ​​the fluid device. In some embodiments, the analyte is trapped within the separate area. In some embodiments, the size and shape of the separate area can be adjusted according to the size, shape, or other properties of the analyte. In some embodiments, an algorithm is used to determine the shape and size of the separate area. In some embodiments, the algorithm is a supervised learning algorithm, a self-supervised learning algorithm, or an unsupervised learning algorithm.

[0031] In some embodiments, the separate area is optically identified. In some embodiments, the detector is configured to detect the location of the analyte within the fluid device. In some embodiments, the detector configured to detect the location of the analyte within the fluid device is a microscope objective lens for imaging the fluid device. In some embodiments, an algorithm is used to determine where the analyte is located based on the imaging. In some embodiments, the imaging is bright-field imaging, phase-contrast imaging, or fluorescence imaging, or any combination thereof. In some embodiments, the algorithm is a supervised learning algorithm, a self-supervised learning algorithm, or an unsupervised learning algorithm. In some embodiments, the objective lens is coupled in the fluid device to an energy source that radiates energy to a separate area.

[0032] In some embodiments, the method further includes the step of introducing one or more reagents into a polymer matrix that reacts with the analyte. In some embodiments, one or more reagents flow through a membrane. In some embodiments, the membrane is semipermeable. In some embodiments, the membrane contains pores. In some embodiments, the pores are less than 10 μm. In some embodiments, one or more reagents include enzymes, drug molecules, oligonucleotides, primers, or any combination thereof. In some embodiments, one or more reagents are dissolving reagents. In some embodiments, one or more reagents are nucleic acid denaturing reagents. In some embodiments, one or more reagents degrade the polymer matrix.

[0033] In some embodiments, the analyte is a cellular component. In some embodiments, the analyte is a small molecule composed of nucleic acids, amino acids, intracellular proteins, surface proteins, secretory proteins, exosomes, metabolites or lipids, or any combination thereof. In some embodiments, the analyte is captured by a capture probe within a polymer matrix. In some embodiments, the analyte is captured by a capture probe on the surface of a polymer matrix.

[0034] In some embodiments, the analyte is released from the cell upon interaction with the reagent. In some embodiments, the reagent is an oxidizing or reducing agent. In some embodiments, the reagent is an organic or inorganic molecule. In some embodiments, the organic or inorganic molecule is a pharmaceutical compound or a surfactant. In some embodiments, the reagent is a protein. In some embodiments, the reagent is a DNA aptamer. In some embodiments, the reagent is a bead containing a biomolecule. In some embodiments, the reagent is a biological species. In some embodiments, the biological species is a virus or a cell.

[0035] In some embodiments, the analyte is released from the cells upon exposure to an energy source. In some embodiments, the energy source is UV light for lysing the cells. In some embodiments, the energy source is visible light for lysing the cells. In some embodiments, UV light is used to activate a photoactivatable surfactant and lyse the cells. In some embodiments, visible light is used to activate a photoactivatable surfactant and lyse the cells.

[0036] In some embodiments, the method further includes the step of identifying an analyte or a derivative thereof. In some embodiments, the analyte is a nucleic acid molecule, and the identification step includes sequencing of the nucleic acid molecule or a derivative thereof. In some embodiments, the method further includes the step of measuring the quality of the analyte. In some embodiments, quality is the shape or size of the analyte.

[0037] In some embodiments, the method further includes the step of performing one or more functional assays to analyze the analytes to evaluate cell viability, cell morphology, cell secretion, cell response, cell-cell interactions, or any combination thereof. In some embodiments, one or more functional assays are colorimetric assays or fluorescence assays. In some embodiments, one or more functional assays are performed using bright-field phase imaging or fluorescence imaging of the analytes.

[0038] In some embodiments, the method further includes the step of performing one or more omics assays to characterize and quantify the analyte or its components. In some embodiments, one or more omics assays are proteomics assays, transcriptomics assays, genomics assays, or epigenomics assays, or any combination thereof. In some embodiments, one or more omics assays are multi-omics assays.

[0039] Another aspect of the present disclosure provides a system comprising a fluid device containing one or more biological components and one or more polymer precursors. The system may further include at least one energy source in communication with the fluid device. In some embodiments, the at least one energy source supplies energy to the fluid device to cause one or more polymer precursors to form at least one polymer matrix on or adjacent to the biological components.

[0040] In some embodiments, the fluid device includes a channel through which it is disposed. In some embodiments, a first surface is disposed along a portion of the channel, and a second surface is disposed opposite to the first surface. In some embodiments, the fluid device includes a chamber disposed within it. In some embodiments, a first surface is disposed along a portion of the chamber, and a second surface is disposed opposite to the first surface. In some embodiments, the first surface is a bottom surface. In some embodiments, the second surface is an top surface. In some embodiments, the fluid device further includes one or more capture elements that immobilize at least one of one or more biological components at a location adjacent to the first surface that forms the immobilized biological components. In some embodiments, the first surface is disposed adjacent to an energy source. In some embodiments, the energy source is a light source. In some embodiments, the energy source is an array of electrodes. In some embodiments, the energy source supplies electrochemical energy to one or more polymer precursors to form an array of polymer matrices. In some embodiments, at least two of the one or more polymer precursors are coupled to the first surface to form a pattern on the first surface.

[0041] In some embodiments, at least one polymer matrix is ​​formed on or adjacent to the pattern. In some embodiments, at least one polymer matrix is ​​coupled to a first surface. In some embodiments, at least one polymer matrix extends from a first surface to a second surface such that at least one polymer matrix surrounds at least a portion of the biological component to which it is immobilized.

[0042] In some embodiments, at least one energy source is in communication with a fluid device via at least one of optical communication, electrochemical communication, electromagnetic communication, thermal communication, or microwave communication. In some embodiments, the at least one energy source includes a photogenerating device, a thermogenerating device, an electrochemical generating device, an electrode, or a microwave device. In some embodiments, the system further includes a photolithography device or a digital micromirror device (DMD) configured to control the spatial distribution of energy from the energy source.

[0043] In some embodiments, one or more capture elements include a physical trap, a geometric trap, a well, an electrochemical trap, a chemical affinity trap, one or more magnetic particles, an electrophoretic trap, a dielectrophoretic trap, or a combination thereof. In some embodiments, the chemical affinity trap includes streptavidin, an antibody, or a combination thereof. In some embodiments, the physical trap includes a polymer matrix. In some embodiments, the polymer matrix includes a hydrogel. In some embodiments, the electrochemical trap includes a gold electrode, a platinum electrode, an indium tin oxide (ITO) electrode, or other suitable electrochemical traps. In some embodiments, one or more capture elements are arranged in a pattern on a first surface. In some embodiments, one or more capture elements include a well. In some embodiments, the diameter of the well is 1 μm (micrometer) to 50 μm. In some embodiments, the depth of the well is 0.1 μm to 100 μm.

[0044] In some embodiments, one or more biological components are multiple biological components. In some embodiments, multiple biological components are coupled to one or more capture elements. In some embodiments, the fluid device is a microfluidic device or a nanofluidic device. In some embodiments, the fluid device is used for nucleic acid sequencing. In some embodiments, nucleic acid sequencing includes next-generation sequencing, short-read sequencing, nanopore sequencing, sequencing by synthesis, sequencing by in situ hybridization, or optical readout.

[0045] In some embodiments, one or more biological components include cells, cell lysates, nucleic acids, microbiomes, proteins, cell mixtures, spatially linked biological components, or metabolites. In some embodiments, the cell mixture includes a first cell type and a second cell type. In some embodiments, the first cell type is different from the second cell type. In some embodiments, the cells are animal cells (e.g., human cells), plant cells, fungal cells, or bacterial cells. In some embodiments, one or more biological components include tumor spheroids or spatially linked biological specimens.

[0046] In some embodiments, the nucleic acid is DNA of 100 base pairs or more, or RNA of 50 bases or more. In some embodiments, the cell lysate contains DNA of 50 bp (base pairs) to 100 Gbp (gigabase pairs), or RNA of 50 bp to 100 kbp (kilobase pairs). In some embodiments, at least one polymer matrix comprises a hydrogel. In some embodiments, the fluid device further comprises one or more polymer precursors. In some embodiments, one or more polymer precursors comprises a hydrogel precursor. In some embodiments, at least one polymer matrix inhibits the passage of immobilized biological components. In some embodiments, at least one polymer matrix forms a polymer matrix wall extending from a first surface to a second surface, thereby forming a chamber within the channel. For example, such a chamber may include a cylinder shell or polygonal shell, comprising an internal space or interior and a polymer matrix wall. In some embodiments, such a chamber has an annular cross-section. As used herein, the term “annular cross-section” means a cross-section that is morphologically equivalent to a ring. In some embodiments, the internal space or interior of the chamber has an inner diameter of 1 μm to 500 μm and a volume in the range of 1 picoliters to 200 nanoliters, or 100 picoliters to 100 nanoliters, or 100 picoliters to 10 nanoliters. In some embodiments, the polymer matrix wall has a thickness of at least 1 μm (micrometer). In some embodiments, the polymer matrix wall having an annular cross-section has an aspect ratio (i.e., height / width) of 1 or less. In some embodiments, the aspect ratio and the thickness of the polymer matrix wall are selected to maximize the stability of the chamber against forces, such as reagent flow through channels, washing, etc. In some embodiments, at least one polymer matrix wall is a hydrogel wall. In some embodiments, at least one polymer matrix is ​​degradable. In some embodiments, the degradation of at least one polymer matrix is ​​"on demand".

[0047] In some embodiments, on-demand degradation may be carried out using polymer precursors that allow photocrosslinking and photodegradation, for example, by using different wavelengths for crosslinking and degradation. For example, eosin Y may be used for radical polymerization in a defined region using a wavelength of 500 nm, and then the crosslinking agent can be cleaved using illumination at 380 nm. In other embodiments, photocage hydrogel cleavage reagents may be included in the formation of polymer matrix walls. For example, a hydrogel can be created using an acid-unstable crosslinking agent (e.g., an ester), and then the hydrogel can be degraded by creating localized acidic conditions using UV light.

[0048] In some embodiments, at least one polymer matrix is ​​degradable by at least one of the following: (i) contacting the at least one polymer matrix with a cleavage reagent; (ii) heating the at least one polymer matrix to at least 90°C; or (iii) exposing the at least one polymer matrix to a wavelength of light that cleaves a photocleavageable crosslinking agent that crosslinks the polymers of the at least one polymer matrix. In some embodiments, the at least one polymer matrix comprises a hydrogel. In some embodiments, the cleavage reagent degrades the hydrogel. In some embodiments, the cleavage reagent comprises a reducing agent, an oxidizing agent, an enzyme, a pH-based cleavage reagent, or a combination thereof. In some embodiments, the cleavage reagent comprises dithiothreitol (DTT), tris(2-carboxyethyl)phosphine (TCEP), tris(3-hydroxypropyl)phosphine (THP), or a combination thereof.

[0049] In some embodiments, at least one polymer matrix allows the passage of a reagent. In some embodiments, at least one polymer matrix contains pores. In some embodiments, the average size of the pores is adjusted using a chemical reagent by applying heat, electricity, light, or a combination thereof. In some embodiments, the reagent includes at least one of an enzyme, a chemical, an oligonucleotide, or a primer having a size of less than 50 base pairs.

[0050] In some embodiments, the reagent comprises lysozyme, proteinase K, random hexamer, polymerase, transposase, ligase, catalytic enzyme, deoxynucleotide triphosphate, buffer, cell culture medium, or divalent cation. In some embodiments, at least one polymer matrix comprises pores that are sized to allow the diffusion of the reagent across at least one polymer matrix, but are too small to allow DNA or RNA for analysis to traverse the pores. In some embodiments, the retained DNA or RNA has a length that can be sequenced using conventional synthetic sequencing techniques. For example, such DNA or RNA comprises at least 50 nucleotides, or in some embodiments, at least 100 nucleotides. In some embodiments, at least one polymer matrix comprises a hydrogel. In some embodiments, the hydrogel is polyethylene glycol (PEG)-thiol, PEG-acrylate, acrylamide, N,N'-bis(acryloyl)cystamine, PEG, polypropylene oxide (PPO), polyacrylic acid, poly(hydroxyethyl methacrylate) (PHEMA), poly(methyl methacrylate) (PMMA), poly(N-isopropylacrylamide) (PNIPAAm), poly(lactic acid) (PLA), poly(lactic acid-co-glycolic acid) (PLGA), polycaprolactone (PCL), poly(vinyl sulfonic acid) (PVSA), poly(L-asphalt) This includes paratic acid, poly(L-glutamic acid), polylysine, agar, agarose, alginate, heparin, alginate sulfate, dextran sulfate, hyaluronan, pectin, carrageenan, gelatin, chitosan, cellulose, collagen, bisacrylamide, diacrylate, diallylamine, triallylamine, divinyl sulfone, diethylene glycol diallyl ether, ethylene glycol diacrylate, polymethylene glycol diacrylate, polyethylene glycol diacrylate, trimethylopropoane trimethacrylate, ethoxylated trimethylol triacrylate, or ethoxylated pentaerythritol tetraacrylate, or combinations or mixtures thereof.In some embodiments, the hydrogel comprises an enzymatically degradable hydrogel, PEG-thiol / PEG-acrylate, acrylamide / N,N'-bis(acryloyl)cystamine (BACy), or PEG / PPO. In some embodiments, a chamber having a degradable polymer matrix (hydrogel) wall can be formed using the following precursors and crosslinkers. Polymer precursors can be formed by using any hydrogel precursor and crosslinker from Table 1A (columns 1 and 3, respectively). The resulting polymer matrix can be degraded by the degraders shown in Table 1A (column 4). [Table 1A] [Table 1B]

[0051] In some embodiments, the first surface, the second surface, or both include one or more barcodes. In some embodiments, one or more barcodes include identifiers for identifying a source of one or more biological components. In some embodiments, the source includes a specimen from which one or more biological components are collected. In some embodiments, the source includes a physiological or anatomical source from which one or more biological components are collected. In some embodiments, the anatomical source includes an organ of interest. In some embodiments, the subject is a human. In some embodiments, one or more barcodes are configured to bind to one or more biological components, or molecules made up of one or more biological components.

[0052] In some embodiments, the first surface, the second surface, or both contain one or more compounds configured to bind to one or more biological components. In some embodiments, the first or second surface is functionalized with a surface polymer. In some embodiments, the surface polymer is functionalized with oligonucleotides, antibodies, cytokines, chemokines, proteins, antibody derivatives, antibody fragments, carbohydrates, toxins, aptamers, or any combination thereof. In some embodiments, the surface of the polymer matrix is ​​functionalized with oligonucleotides, antibodies, cytokines, chemokines, proteins, antibody derivatives, antibody fragments, carbohydrates, toxins, aptamers, or any combination thereof. In some embodiments, the surface polymer includes polyethylene glycol (PEG), silane polymers, pyridine carboxyaldehyde (PCA), acrylamide, agarose, or a combination thereof.

[0053] In some embodiments, the system further includes a detector for identifying one or more biological components, one or more barcodes, or combinations thereof. In some embodiments, the detector includes a camera (fluorescence camera).

[0054] In some embodiments, the system further includes a stage for holding a fluid device. In some embodiments, the system further includes a sequencing device for obtaining sequencing data. In some embodiments, the sequencing data is generated using next-generation sequencing, short-read sequencing, nanopore sequencing, sequencing by synthesis, sequencing by in situ hybridization, or optical readout.

[0055] In some embodiments, the system further includes a spatial energy modulator that selectively supplies energy to a fluid device. In some embodiments, the spatial energy modulator is generated using a detector that identifies the location of at least one biological component. In some embodiments, the spatial energy modulator includes a physical photomask, a virtual photomask, a physical electrode distribution pattern, or a virtual electrode distribution pattern. In some embodiments, the spatial energy modulator includes a photolithography mask or a digital micromirror device (DMD) mask.

[0056] Another aspect of the present disclosure provides a method for analyzing biological components. The method may include (a) introducing one or more biological components into a fluid device; (b) positioning a first portion of one or more biological components adjacent to a first surface of the fluid device; and (c) forming one or more polymer matrices adjacent to the first portion of the first surface to localize at least one of the one or more biological components to the first portion.

[0057] In some embodiments, the method further includes (d) stirring one or more biological components within a fluid device, (e) positioning a second portion of one or more biological components adjacent to a first surface of the fluid device, and (f) forming one or more polymer matrices adjacent to the second portion of the first surface to immobilize at least one of the one or more biological components of the second portion. The stirring step in this and other embodiments can prevent or disrupt the aggregation of biological components, such as living cells. Stirring can also induce a more uniform distribution of the biological components on the first surface within the channel.

[0058] In some embodiments, the method further includes the step of locating at least one of one or more biological components such that at least one energy source supplies energy to a fluid device to form one or more polymer matrices on or adjacent to a identified location.

[0059] Another aspect of the present disclosure provides a method for analyzing biological components. The method may include (a) introducing a biological component into a fluid device; (b) coupling the biological component to one or more capture elements disposed on a first or second surface of the fluid device to obtain a coupled biological component; and (c) forming a polymer matrix on or adjacent to the coupled biological component.

[0060] In some embodiments, the method further includes the step of introducing one or more polymer precursors into a fluid device. In some embodiments, the step of forming a polymer matrix includes supplying energy to the fluid device to form the polymer matrix.

[0061] In some embodiments, energy is selectively supplied to one or more parts of a fluid device. In some embodiments, the method further includes a spatial energy modulating element that selectively supplies energy to the fluid device. In some embodiments, the spatial energy modulating element includes a physical photomask, a virtual photomask, a physical electrode distribution pattern, or a virtual electrode distribution pattern.

[0062] In some embodiments, the spatial energy adjustment element includes a photolithography mask or a digital micromirror device (DMD) mask. In some embodiments, energy is supplied via a light energy source, a thermal energy source, an electrochemical energy source, or an electromagnetic energy source.

[0063] In some embodiments, the polymer matrix is ​​coupled to a first surface. In some embodiments, energy forms the polymer matrix around a portion of the coupled biological component. In some embodiments, at least a portion of the biological component is encapsulated by the polymer matrix. In some embodiments, the entire biological component is encapsulated by the polymer matrix.

[0064] In some embodiments, the method further includes the steps of coupling a first biological component to a first capture element to form a first analysis chamber, and coupling a second biological component to a second capture element to form a second analysis chamber. In some embodiments, the first analysis chamber is adjacent to the second analysis chamber. In some embodiments, the first analysis chamber is located 5 micrometers (μm) to 1,000 μm away from the second analysis chamber.

[0065] In some embodiments, the method further includes the steps of analyzing a first biological component in a first analysis chamber and analyzing a second biological component in a second analysis chamber. In some embodiments, the method further includes the steps of initiating a first reaction in the first biological component and initiating a second reaction in the second biological component. In some embodiments, the first and second reactions are different. In some embodiments, the method further includes the steps of initiating a third reaction in the first biological component and initiating a fourth reaction in the second biological component. In some embodiments, the third and fourth reactions are different.

[0066] In some embodiments, the method further includes the step of obtaining a genome, transcriptome, proteome, epigenome, methylome, secretome, or metabolome of coupled biological components.

[0067] In some embodiments, the proteome includes secreted proteins, surface proteins, or a combination thereof. In some embodiments, the transcriptome is substantially a full-length transcriptome. In some embodiments, the transcriptome is a full-length transcriptome. In some embodiments, the method further includes the step of sequencing at least one nucleic acid of the biological components. In some embodiments, the sequencing step does not include amplification of the sequencing library. In some embodiments, the nucleic acid library derived from the biological components is sequenced in the same chamber. In some embodiments, the method further includes the step of coupling a barcode to the biological component or a molecule produced by the biological component.

[0068] In some embodiments, the method further includes the step of exposing the analyte to a biological component or coupled biological component. In some embodiments, the biological component comprises one or more microorganisms. In some embodiments, the analyte comprises an antimicrobial agent or a microbial growth promoter. In some embodiments, the method further includes the step of screening one or more microorganisms for susceptibility to the antimicrobial agent. In some embodiments, the analyte comprises a pharmaceutical agent.

[0069] In some embodiments, the method further includes a step of screening the effects of a pharmaceutical agent on biological components. In some embodiments, the method further includes a step of screening biological components for the production of a target molecule.

[0070] In some embodiments, the target molecule includes at least one of the following: an antibody, cytokine, chemokine, protein, antibody derivative, antibody fragment, carbohydrate, toxin, or aptamer.

[0071] In some embodiments, the method further includes the step of forming a polymer matrix around a biological component such that the biological component is positioned within a structure formed by the polymer matrix.

[0072] In some embodiments, the method further includes the step of analyzing local parameters in a first or second analysis chamber. In some embodiments, the level of the local parameter in the first analysis chamber is different from the level of the local parameter in the second analysis chamber. In some embodiments, the local parameter includes pH, oxygen concentration, or CO2 concentration.

[0073] In some embodiments, one or more capture elements include a polymer matrix. In some embodiments, the polymer matrix includes a hydrogel.

[0074] Another aspect of the present disclosure provides a method for obtaining a transcriptome of a biological component. The method may include (a) forming a polymer matrix on or adjacent to the biological component to form an analysis chamber, and (b) carrying out one or more reactions in the analysis chamber to obtain a transcriptome of the biological component. In some embodiments, the biological component remains in or substantially in the analysis chamber during the carrying out of one or more reactions.

[0075] In some embodiments, the method further includes the step of coupling biological components to capture elements placed in a fluid device to obtain coupled biological components. In some embodiments, the method further includes the step of supplying energy from an energy source to a fluid device to form a polymer matrix. In some embodiments, the energy is supplied using a spatial energy modulating element selectively. In some embodiments, the spatial energy modulating element is generated based on the location of the biological components. In some embodiments, the spatial energy modulating element includes a physical photomask, a virtual photomask, a physical electrode distribution pattern, a virtual electrode distribution pattern, a photolithography mask, or a digital micromirror device (DMD) mask.

[0076] In some embodiments, the biological components include RNA. In some embodiments, the RNA is 50 to 100 kb (kilobase-based). In some embodiments, the polymer matrix includes pores that are sized to allow the diffusion of reagents across the polymer matrix, but are too small to allow RNA to traverse the pores. In some embodiments, one or more reactions include RNA sequencing.

[0077] Another aspect of the present disclosure provides a method for analyzing two or more biological components. The method may include (a) introducing a first biological component and a second biological component into a fluid device; (b) forming a polymer matrix on or adjacent to the first biological component to form a first analysis chamber, and forming a polymer matrix on or adjacent to the second biological component to form a second analysis chamber; and (c) analyzing one or more properties of the first biological component and the second biological component. In some embodiments, the first analysis chamber is adjacent to the second analysis chamber in the fluid device. In some embodiments, one or more properties include a first property and a second property. In some embodiments, (c) includes analyzing a first property and a second property of the first biological component in the first analysis chamber.

[0078] In some embodiments, the first biological component remains in the first analysis chamber during the analysis of the first and second properties, respectively. In some embodiments, one or more properties include a response to an analyte, a response to a pharmaceutical agent, a response to an antimicrobial agent, the production of a target compound, the production of a target molecule, the production of a nucleic acid, or the production of a protein. In some embodiments, the first biological component is biologically in communication with the second biological component. In some embodiments, the biological communication generates a biological response in the first or second biological component. In some embodiments, the biological communication includes molecules, such as proteins, nucleic acids, cytokines, chemokines, or combinations thereof, produced by the first or second biological component.

[0079] Another aspect of this disclosure provides a method for identifying nucleic acid molecules. The method may include the steps of providing a polymer matrix containing nucleic acid molecules and detecting nucleic acid molecules in the absence of nucleic acid amplification.

[0080] In some embodiments, the nucleic acid molecule is a deoxyribonucleic acid (DNA) molecule.

[0081] In some embodiments, the polymer matrix forms chambers for localizing nucleic acids. In some embodiments, the chambers are formed on demand. In some embodiments, the polymer matrix is ​​degraded on demand. In some embodiments, the DNA is 100 base pairs or larger. In some embodiments, the nucleic acid is a ribonucleic acid molecule (RNA). In some embodiments, the RNA is 50 nucleotides or larger. In some embodiments, the method further includes the step of generating a nucleic acid library from biological components within the chambers. In some embodiments, the nucleic acid library is sequenced within the chambers.

[0082] Another aspect of this disclosure provides a method for processing biological components. The method may include the step of determining a genome sequence, transcriptome, proteome, or epigenome in the absence of nucleic acid amplification. In some embodiments, the processing is carried out in a single microfluidic device. In some embodiments, the cells are at least partially contained within a polymer matrix. In some embodiments, the polymer matrix is ​​degraded on demand. In some embodiments, the method further includes the step of determining methylation in the cells.

[0083] Another aspect of this disclosure provides a method comprising the step of identifying multiple nucleic acid molecules in multiple cells without barcoding the individual nucleic acid molecules of the multiple nucleic acid molecules. In some embodiments, the steps of extracting and identifying the multiple nucleic acid molecules are performed in a single microfluidic device. In some embodiments, the identifying step includes sequencing. In some embodiments, the method further includes the step of forming a polymer matrix on or adjacent to individual cells of the multiple cells so that the individual cells are separated from each other. In some embodiments, the method further includes the step of extracting individual nucleic acid molecules from individual cells. In some embodiments, the sequencing includes sequencing of individual nucleic acid molecules in the polymer matrix. In some embodiments, the sequencing includes next-generation sequencing, short-read sequencing, nanopore sequencing, sequencing by synthesis, sequencing by in situ hybridization, or optical readout.

[0084] Another aspect of the present disclosure provides a method comprising the steps of (a) providing a plurality of nucleic acid molecules in a plurality of matrices, and (b) sequencing the plurality of nucleic acid molecules while they are in the plurality of matrices. In some embodiments, the individual nucleic acid molecules of the plurality of nucleic acid molecules are derived from different cells. In some embodiments, the plurality of matrices are arranged in a fluid device. In some embodiments, the plurality of matrices contain a plurality of cells. In some embodiments, the plurality of cells contain a plurality of nucleic acid molecules. In some embodiments, the sequencing includes next-generation sequencing, short-read sequencing, nanopore sequencing, sequencing by synthesis, sequencing by in situ hybridization, or optical readout.

[0085] Another aspect of the present disclosure provides a method for analyzing biological components. The method may include (a) introducing one or more biological components into a fluid device; (b) positioning a first portion of one or more biological components adjacent to a first surface of the fluid device; and (c) forming one or more polymer matrices adjacent to the first portion of the first surface to localize at least one of the one or more biological components to the first portion. In some embodiments, the method further includes (d) stirring one or more biological components within the fluid device; (e) positioning a second portion of one or more biological components adjacent to a first surface of the fluid device; and (f) forming one or more polymer matrices adjacent to the second portion of the first surface to immobilize at least one of the one or more biological components of the second portion. In some embodiments, the method further includes a step of locating at least one of the one or more biological components such that at least one energy source supplies energy to the fluid device to form one or more polymer matrices on or adjacent to a specified location.

[0086] Another aspect of the present disclosure provides a system including a fluid device. The fluid device may include a flow channel, an analysis channel located adjacent to the flow channel, a layer located between the flow channel and the analysis channel, and at least one energy source communicating with the fluid device. In some embodiments, at least one flow-inhibiting element is located within the flow channel to obstruct the flow of biological components. In some embodiments, the layer includes at least one sealable opening located adjacent to the at least one flow-inhibiting element. In some embodiments, the at least one sealable opening is configured to allow the passage of biological components. In some embodiments, at least one energy source is configured to form a polymer matrix within the analysis channel.

[0087] In some embodiments, a portion of the flow channel is substantially parallel to a portion of the analysis channel. In some embodiments, at least one sealable opening is configured to transition from a sealed state to an open state. In some embodiments, the passage of biological components through at least one sealable opening is inhibited in the sealed state. In some embodiments, when at least one sealable opening is in a sealed state, at least one sealable opening is sealed with at least one of agarose gel, a temperature-soluble polymer, an N-isopropylacrylamide (NIPAAm) polymer, a wax compound, or an alginate.

[0088] In some embodiments, the flow channel includes a surface positioned opposite the flow channel surface of the layer. In some embodiments, at least one of the at least one inhibitory element extends from the surface toward the flow channel surface such that the flow of biological components in the flow channel is inhibited by the at least one inhibitory element. In some embodiments, the analysis channel includes a surface positioned opposite the analysis channel surface of the layer. In some embodiments, the flow channel is removablely coupled to the analysis channel. In some embodiments, the surface of the analysis channel includes one or more barcodes. In some embodiments, the barcodes include oligonucleotides.

[0089] In some embodiments, the polymer matrix is ​​coupled to the surface of the analytical channel or to at least one of the analytical channel surfaces of the layer. In some embodiments, the polymer matrix extends from the surface of the analytical channel to the analytical channel surface of the layer such that the polymer matrix surrounds at least a portion of the biological components.

[0090] In some embodiments, at least one energy source is in communication with a fluid device by at least one of optical, electrochemical, electromagnetic, thermal, or microwave communication. In some embodiments, the at least one energy source includes a photogenerating device, a thermogenerating device, an electrochemical generating device, an electrode, or a microwave device.

[0091] In some embodiments, the biological component comprises multiple biological components.

[0092] In some embodiments, the fluid device is a microfluidic device or a nanofluidic device. In some embodiments, the fluid device includes a sequencing flow cell. In some embodiments, the fluid device is used for nucleic acid sequencing. In some embodiments, the biological components include cells, nucleic acids, microbiomes, proteins, combinations of cells, spatially linked biological components, or metabolites. In some embodiments, cells are animal cells (e.g., human cells), plant cells, fungal cells, bacterial cells, tumor spheroids, or combinations thereof. In some embodiments, nucleic acids are DNA with 100 base pairs or more, or RNA with 50 base pairs or more.

[0093] In some embodiments, the polymer matrix comprises a hydrogel. In some embodiments, the fluid device further comprises one or more polymer precursors. In some embodiments, one or more polymer precursors comprises a hydrogel precursor. In some embodiments, the polymer matrix comprises polymer matrix walls having a width of at least 1 μm. In some embodiments, the polymer matrix inhibits the passage of biological components. In some embodiments, the polymer matrix walls are hydrogel walls. In some embodiments, the polymer matrix is ​​degradable. In some embodiments, the degradation of the polymer matrix is ​​"on demand". In some embodiments, the polymer matrix is ​​degradable by at least one of (i) contacting the polymer matrix with a cleavage reagent, (ii) heating the polymer matrix to at least 90°C, or (iii) exposing the polymer matrix to a wavelength of light that cleaves a photocleavageable crosslinking agent that crosslinks the polymers of the polymer matrix.

[0094] In some embodiments, a sealable opening is transitioned to an open state by at least one of the following: (i) contacting the sealable opening with a cleavage reagent; (ii) heating the sealable opening to at least 90°C; or (iii) exposing the sealable opening to a wavelength of light that cleaves a photocleavageable crosslinking agent that crosslinks the polymer of the sealable opening. In some embodiments, the polymer matrix comprises a hydrogel. In some embodiments, the cleavage reagent is configured to decompose the polymer matrix. In some embodiments, the cleavage reagent comprises a reducing agent, an oxidizing agent, an enzyme, a pH-based cleavage reagent, or a combination thereof. In some embodiments, the cleavage reagent comprises dithiothreitol (DTT), tris(2-carboxyethyl)phosphine (TCEP), tris(3-hydroxypropyl)phosphine (THP), or a combination thereof.

[0095] In some embodiments, the polymer matrix allows the passage of reagents. In some embodiments, the polymer matrix includes pores. In some embodiments, the size of the pores is controlled by changing the composition of one or more polymer precursors, at least one energy source, or a combination thereof.

[0096] In some embodiments, the reagent comprises at least one of the following: an enzyme, a chemical, an oligonucleotide, or a primer having a size of less than 50 base pairs. In some embodiments, the reagent comprises lysozyme, proteinase K, random hexamer, polymerase, transposase, ligase, catalytic enzyme, deoxynucleotide triphosphate, buffer, cell culture medium, or divalent cation.

[0097] In some embodiments, the polymer matrix contains pores that are sized to allow the diffusion of reagents across the matrix but are too small to allow DNA or RNA to traverse the pores. In some embodiments, the polymer matrix comprises a hydrogel. In some embodiments, the hydrogel comprises polyethylene glycol (PEG)-thiol, PEG-acrylate, acrylamide, N,N'-bis(acryloyl)cystamine, PEG, polypropylene oxide (PPO), polyacrylic acid, poly(hydroxyethyl methacrylate) (PHEMA), poly(methyl methacrylate) (PMMA), poly(N-isopropylacrylamide) (PNIPAAm), poly(lactic acid) (PLA), poly(lactic acid-co-glycolic acid) (PLGA), polycaprolactone (PCL), poly(vinyl sulfonic acid) (PVSA), poly(L-asulfonic acid) The hydrogels include paratic acid, poly(L-glutamic acid), polylysine, agar, agarose, alginate, heparin, alginate sulfate, dextran sulfate, hyaluronan, pectin, carrageenan, gelatin, chitosan, cellulose, collagen, bisacrylamide, diacrylate, diallylamine, triallylamine, divinyl sulfone, diethylene glycol diallyl ether, ethylene glycol diacrylate, polymethylene glycol diacrylate, polyethylene glycol diacrylate, trimethylopropoane trimethacrylate, ethoxylated trimethylol triacrylate, or ethoxylated pentaerythritol tetraacrylate, or combinations or mixtures thereof. In some embodiments, the hydrogels include enzymatically digestible hydrogels, PEG-thiol / PEG-acrylate, acrylamide / N,N'-bis(acryloyl)cystamine (BACy), or PEG / PPO.

[0098] In some embodiments, the surface includes one or more barcodes. In some embodiments, the surface of the analytical channel includes one or more compounds configured to bind to biological components. In some embodiments, the surface of the analytical channel is functionalized with a surface polymer. In some embodiments, the surface polymer is functionalized with oligonucleotides, antibodies, cytokines, chemokines, proteins, antibody derivatives, antibody fragments, carbohydrates, toxins, aptamers, or any combination thereof. In some embodiments, the surface of the polymer matrix is ​​functionalized with oligonucleotides, antibodies, cytokines, chemokines, proteins, antibody derivatives, antibody fragments, carbohydrates, toxins, aptamers, or any combination thereof. In some embodiments, the surface polymer includes polyethylene glycol (PEG), silane polymers, pyridinecarboxaldehyde (PCA), acrylamide, agarose, or a combination thereof.

[0099] In some embodiments, the system further includes a detector for identifying one or more biological components, one or more barcodes, or combinations thereof. In some embodiments, the detector includes a camera. In some embodiments, the system further includes a stage for holding a fluid device. In some embodiments, the system further includes a sequencing device for obtaining sequencing data.

[0100] In some embodiments, the system further includes a spatial energy modulating element that selectively supplies energy to a fluid device. In some embodiments, the spatial energy modulating element includes a physical photomask, a virtual photomask, a physical electrode distribution pattern, or a virtual electrode distribution pattern. In some embodiments, the spatial energy modulating element includes a photolithography mask or a digital micromirror device (DMD) mask.

[0101] Another aspect of this disclosure provides a method for analyzing biological components. The method may include (a) introducing biological components into a flow channel of a fluid device; (b) inhibiting the flow of biological components adjacent to an inhibitory element; (c) positioning the biological components from the flow channel of the fluid device to an analysis channel; and (d) forming a polymer matrix on or adjacent to the biological components in the analysis channel, either before or after positioning the biological components in the analysis channel. In some embodiments, a sealable opening is positioned adjacent to the inhibitory element.

[0102] In some embodiments, prior to the placement step in (c), the sealable opening is decomposed by at least one of the following: (i) contacting the sealable opening with a cutting reagent; (ii) heating the sealable opening to at least 90°C; or (iii) exposing the sealable opening to a wavelength of light that cuts a photocutting crosslinking agent that crosslinks the polymer of the sealable opening. In some embodiments, the method further includes the step of introducing one or more polymer precursors into a fluid device. In some embodiments, the step of forming a polymer matrix includes supplying energy to the fluid device to form a polymer matrix. In some embodiments, the energy is supplied selectively to one or more parts of the fluid device. In some embodiments, the method further includes the step of activating a spatial energy modulating element to selectively supply energy to the fluid device. In some embodiments, the spatial energy modulating element includes a physical photomask, a virtual photomask, a physical electrode distribution pattern, or a virtual electrode distribution pattern. In some embodiments, the spatial energy modulating element includes a photolithography mask or a digital micromirror device (DMD) mask. In some embodiments, energy is supplied via a light energy source, a thermal energy source, an electrochemical energy source, or an electromagnetic energy source.

[0103] In some embodiments, the polymer matrix is ​​coupled to the surface of the analytical channel. In some embodiments, the energy forms a polymer matrix around a portion of the coupled biological component. In some embodiments, at least a portion of the biological component is encapsulated by the polymer matrix. In some embodiments, the entire biological component is encapsulated by the polymer matrix.

[0104] In some embodiments, the method further includes the steps of encapsulating a first biological component to form a first analysis chamber, and encapsulating a second biological component to form a second analysis chamber. In some embodiments, the first analysis chamber is adjacent to the second analysis chamber. In some embodiments, the first analysis chamber is located 5 micrometers (μm) to 1,000 μm away from the second analysis chamber.

[0105] In some embodiments, the method further includes the steps of analyzing a first biological component in a first analysis chamber and analyzing a second biological component in a second analysis chamber. In some embodiments, the method further includes the steps of initiating a first reaction in the first biological component and initiating a second reaction in the second biological component. In some embodiments, the first and second reactions are different. In some embodiments, the method further includes the steps of initiating a third reaction in the first biological component and initiating a fourth reaction in the second biological component. In some embodiments, the third and fourth reactions are different.

[0106] In some embodiments, the method further includes the step of obtaining a genome, transcriptome, proteome, epigenome, methylome, secretome, or metabolome of a biological component. In some embodiments, the transcriptome is substantially a full-length transcriptome. In some embodiments, the transcriptome is a full-length transcriptome. In some embodiments, the method further includes the step of sequencing the biological component. In some embodiments, the sequencing step does not include amplification of the sequencing library. In some embodiments, the method further includes a barcode configured to be coupled to the biological component or a molecule produced by the biological component.

[0107] In some embodiments, the method further includes the step of exposing the analyte to a biological component or a first biological component. In some embodiments, the biological component comprises one or more microorganisms. In some embodiments, the analyte comprises an antimicrobial agent, a microbial growth-promoting chemical, or a combination thereof. In some embodiments, the method further includes the step of screening one or more microorganisms for susceptibility to the antimicrobial agent. In some embodiments, the analyte comprises a pharmaceutical agent.

[0108] In some embodiments, the method further includes a step of screening the effects of a pharmaceutical agent on biological components. In some embodiments, the method further includes a step of screening biological components for the production of a target molecule. In some embodiments, the target molecule includes at least one of an antibody, cytokine, chemokine, protein, antibody derivative, antibody fragment, carbohydrate, toxin, or aptamer.

[0109] In some embodiments, the method further includes the step of forming a polymer matrix around a biological component such that the biological component is placed within a structure formed by the polymer matrix. In some embodiments, the method further includes the step of analyzing local parameters in a first or second analysis chamber. In some embodiments, the level of the local parameter in the first analysis chamber is different from the level of the local parameter in the second analysis chamber. In some embodiments, the local parameter includes pH, oxygen concentration, or CO2 concentration.

[0110] Another aspect of this disclosure provides a non-temporary computer-readable medium containing machine-executable code that, when executed by one or more computer processors, performs any of the methods described above or elsewhere in this specification.

[0111] In some embodiments, the present invention provides a method for analyzing biological components of a biological sample, such as one or more mammalian cells, comprising the steps of: (a) (i) a channel having an inlet and an outlet, wherein the channel is bounded by a first wall having a first surface and a second wall having a second surface, and the first and second walls are arranged facing each other across the channel; (ii) a fluid device having a spatial energy modulator positioned adjacent to the first wall, the spatial energy modulator capable of projecting energy having predetermined beam characteristics across the channel; (b) loading a reaction mixture comprising a biological sample and one or more polymer precursors into the channel; and (c) synthesizing one or more chambers in the channel around each of the one or more biological components by projecting energy across the channel such that the projected energy crosslinks one or more polymer precursors to form polymer matrix walls of chambers. In some embodiments, the channel is a plane with a shape having a large width and breadth relative to its height. The cross-section of the channel perpendicular to the direction of reagent flow is typically rectangular. The reaction mixture, or other reagents, such as a washing solution, is loaded into the channel through the inlet and removed through the outlet. In some embodiments, such reagents or mixtures are pumped into the channel through the inlet, for example, using a syringe pump. In some embodiments, the outlet may be a vent port to release air from the channel as soon as the reagent is loaded into the channel through the inlet, thereby replacing the air.

[0112] In some embodiments, the present invention provides a method for analyzing biological components of a biological sample, such as one or more mammalian cells, comprising the steps of: (a) (i) a channel, wherein the channel is bounded by a first wall having a first surface; (ii) a spatial energy modulator positioned adjacent to the first wall, wherein the spatial energy modulator can project energy having predetermined beam characteristics into the channel across the first wall; (b) loading a reaction mixture into the channel, comprising the steps of: (c) synthesizing one or more chambers in the channel around each of the one or more biological components by projecting energy into the channel such that the projected energy crosslinks one or more polymer precursors to form polymer matrix walls of chambers. This embodiment may be referred to herein as an “open configuration” of the system of the present invention. In some embodiments of the open configuration, the chambers include a well or container with an open top. In some embodiments, the top of the well or container faces the first surface.

[0113] In some embodiments, the fluid device of the method further includes a detector positioned adjacent to the second wall or opposite the first wall from the spatial energy modulating element in an open configuration embodiment. The detector is positioned to detect optical signals from one or more biological components, e.g., mammalian cells, distributed across the first surface in the chamber. In some embodiments, the first and second walls each include an optically transparent material, for example, the spatial energy modulating element may project light energy into the channel, and as a result, the detector may detect optical signals, e.g., fluorescence or reflected light from the biological components. In some embodiments, the energy projected from the spatial energy modulating element is light energy from a light beam. In some embodiments, the light beam projected by the spatial energy modulating element may have a complex cross-section (in various embodiments) that allows for the simultaneous synthesis of multiple chambers. Examples of optically transparent materials, but not limited to, include glass, quartz, plastics, and similar materials.

[0114] In some embodiments, the step of synthesizing a chamber includes positioning the chamber to encapsulate one or more biological components based on an optical signal detected by a detector. That is, in some embodiments, the detector is operationally related with a spatial energy modulator to selectively project one or more light beams to a location where the detected optical signal indicates the presence of the biological component of interest. In such embodiments, the detector and the spatial energy modulator are operationally related so that the spatial energy modulator is configured to produce an energy beam having a predetermined beam feature. For example, one such feature may be a cross-section of the beam that brings the biological component of interest to be encapsulated in the chamber. In such operational association, the optical signal detected by the detector may include, but are not limited to, the morphology of the biological component, e.g., cell morphology; the motility of the biological component, e.g., cell motility; interactions between one cell type and another, e.g., binding of one cell type to another; the presence, absence, or amount of a label on the biological component.

[0115] In some embodiments, when a chamber is formed or synthesized, the polymer matrix wall of the chamber extends from a first surface to a second surface, forming chambers having interiors. In other embodiments, the fluid device may not have a second wall having a second surface, which is referred to herein as an “open configuration”. In such embodiments, the chamber forms an open cylinder or well shape on the first surface. The height and thickness of the well wall are partially determined by the intensity and duration of light irradiation by a space energy modulating element. In some embodiments, a given beam feature includes a beam having an annular cross-section. In other embodiments, a given beam feature includes a beam having a cross-section including a plurality of annular shapes, which may be separated and discontinuous or include continuous annular shapes. In other embodiments, for example, as shown in Figure 25B, a given beam feature includes a beam that generates a plurality of chambers, where a portion of the chambers shares a polymer matrix wall with one or more other chambers of the plurality.

[0116] In some embodiments, the loading step may further include a step of stirring the reaction mixture, for example, by vibrating, shaking, or stirring the channel, to reduce aggregation of biological components. Such stirring may last for several minutes, for example, 1 to 30 minutes, or for 1 hour or longer, for example, 1 to 2 hours.

[0117] In some embodiments, a first or second surface, typically the first surface, includes one or more capture elements for specifically capturing one or more predetermined biological components of the biological sample, e.g., selected cells, e.g., lymphocytes. In some embodiments, the loading step may include incubating a reaction mixture containing such predetermined biological components under conditions that allow one or more capture elements to capture one or more predetermined biological components. Such incubation may range from a few minutes to an hour or longer. In some embodiments, such incubation time may range from one minute to 10 hours, or from 10 minutes to 2 hours, or from 30 minutes to 2 hours.

[0118] In some embodiments, the method of the present invention includes a step of removing the reaction mixture from the channel after the step of synthesizing the chamber is completed. Such a removal step may include washing the channel with a buffer solution.

[0119] In some embodiments, the method of the present invention includes the step of loading an analytical assay reagent into the channel after the chamber has been formed, for determining one or more features of the biological components enclosed in the chamber, wherein the analytical assay reagent can pass through the polymer matrix wall and generate one or more optical signals indicating one or more features. A wide variety of analytical assays may be performed, including RNA or DNA identification and / or sequencing, protein identification and / or quantification, omics assays, and the like, as shown elsewhere in this application.

[0120] In some embodiments, the polymer matrix wall of the chamber is degradable by treating the chamber with a degrading agent. In such embodiments, the method of the present invention may include the steps of (a) identifying one or more of the chambers having the biological components having selected characteristics based on the one or more optical signals generated from the manipulation of an analytical assay reagent; (b) loading a second reaction mixture containing a second polymer precursor into a channel, wherein the second polymer precursor can form a second polymer matrix wall that is not degradable to at least one degrading agent; and (c) synthesizing a second chamber for enclosing the identified chamber. In some such embodiments, the original chamber having a degradable polymer matrix wall may be degraded by treatment with a degrading agent that leaves the second chamber intact, allowing the biological components of interest to be isolated.

[0121] Another aspect of this disclosure provides a system comprising one or more computer processors and computer memory connected thereto. The computer memory contains machine-executable code that, when executed by one or more computer processors, implements any of the methods described above or elsewhere in this specification.

[0122] Additional aspects and advantages of the present disclosure will be readily apparent to those skilled in the art from the following detailed description, which shows and describes only exemplary embodiments of the present disclosure. As realized, other and different embodiments are possible, and some of their details can be modified in various obvious ways without departing entirely from the present disclosure. Accordingly, the drawings and description should be considered illustrative in nature and not restrictive. In embodiments of the present invention, for example, the following items are provided. (Item 1) A method for processing analytes, (a) (i) providing a fluid device comprising the analyte and (ii) one or more polymer precursors, (b) the step of identifying a separate area within the fluid device, (c) A step of selectively supplying units of energy generated from an energy source to the fluid device to generate a polymer matrix from the one or more polymer precursors within the fluid device. Includes, The polymer matrix is ​​located within the separate area in (b) or adjacent to the separate area in (b), method. (Item 2) The method according to item 1, wherein the fluid device includes a flow channel. (Item 3) The method according to item 1, wherein the fluid device includes an open configuration. (Item 4) The method according to item 1, wherein the fluid device includes one or more distinct locations, and the one or more distinct locations are not in fluid communication with another distinct location. (Item 5) The method according to item 4, wherein the one or more distinct locations are one or more well plates. (Item 6) The method according to item 4, wherein one or more of the aforementioned locations are open at the top. (Item 7) The method according to item 4, wherein the one or more separate locations contain the analyte. (Item 8) The method according to item 1, wherein the fluid device further comprises one or more monomers. (Item 9) The method according to item 1, wherein the fluid device further includes a spatial energy regulating element. (Item 10) The method according to item 1, wherein the fluid device includes a surface on which a capture probe is immobilized, and the capture probe is coupled to the analyte to immobilize the analyte on the surface. (Item 11) The method according to item 1, wherein the polymer matrix is ​​generated adjacent to or surrounding the analyte to immobilize the analyte. (Item 12) The method according to item 1, wherein the polymer matrix forms a hydrogel. (Item 13) The method according to item 10, wherein the capture probe comprises one or more functional groups that can interact with the analyte. (Item 14) The method according to item 13, wherein the one or more functional groups include a DNA sequence complementary to the target DNA or RNA. (Item 15) The method according to item 1, wherein the energy source is in optical communication with the fluid device. (Item 16) The method according to item 1, wherein the energy source is a photogenerating device. (Item 17) The method according to item 16, wherein the light generation device generates light in the range of 350 nm to 800 nm. (Item 18) The method according to item 16, wherein the light generation device generates light in the range of 350 nm to 600 nm. (Item 19) The method according to item 16, wherein the light generation device generates light in the range of 350 nm to 450 nm. (Item 20) The method according to item 16, wherein the light generation device generates UV light. (Item 21) (d) The method according to item 1, wherein the method is performed using a spatial light modulator (SLM). (Item 22) The method according to item 21, wherein the SLM is a digital micromirror device (DMD). (Item 23) The method according to item 21, wherein the SLM is a laser beam guided using a galvanometer. (Item 24) The method according to item 21, wherein the SLM is liquid crystal based. (Item 25) The method according to item 1, wherein the area of ​​the separate area is less than the area of ​​the fluid device. (Item 26) The method according to item 1, wherein the aforementioned separate area contains the analyte. (Item 27) The method according to item 1, wherein the analyte is captured within the separate area. (Item 28) The method according to item 1, wherein the size and shape of the separate areas are adjustable according to the size of the analyte, the shape of the analyte, or other properties of the analyte. (Item 29) The method according to item 28, wherein an algorithm is used to determine the shape and size of the separate areas. (Item 30) The method according to item 29, wherein the algorithm is a supervised learning algorithm, a self-supervised learning algorithm, or an unsupervised learning algorithm. (Item 31) The method according to item 1, wherein the identifying step in (b) is performed optically. (Item 32) The method according to item 1, wherein the identifying step in (b) is performed using a detector configured to detect the location of the analyte within the fluid device. (Item 33) The method according to item 32, wherein the detector configured to detect the location of the analyte within the fluid device is a microscope objective lens for imaging the fluid device. (Item 34) Using the algorithm, the location of the analyte is determined based on the imaging. The method described in item 33 for determining whether or not. (Item 35) The method according to item 33, wherein the imaging is bright-field imaging, phase-contrast imaging, or fluorescence imaging, or any combination thereof. (Item 36) The method according to item 34, wherein the algorithm is a supervised learning algorithm, a self-supervised learning algorithm, or an unsupervised learning algorithm. (Item 37) The method according to item 33, wherein the objective lens is coupled to an energy source that radiates energy to the separate area in the fluid device. (Item 38) The method according to item 1, further comprising the step of introducing one or more reagents into the polymer matrix that reacts with the analyte. (Item 39) The method according to item 38, wherein one or more of the reagents flow through a membrane. (Item 40) The method according to item 39, wherein the film is semipermeable. (Item 41) The method according to item 39, wherein the membrane includes pores. (Item 42) The method according to item 41, wherein the pore size is less than 10 μm. (Item 43) The method according to item 38, wherein the one or more reagents are an enzyme, a drug molecule, an oligonucleotide, a primer, or any combination thereof. (Item 44) The method according to item 38, wherein one or more of the reagents are dissolving reagents. (Item 45) The method according to item 38, wherein one or more of the reagents are nucleic acid denaturation reagents. (Item 46) The method according to item 38, wherein one or more reagents degrade the polymer matrix. (Item 47) The method according to item 1, wherein the analyte is a component of a cell. (Item 48) The method according to item 47, wherein the analyte is a small molecule composed of nucleic acids, amino acids, intracellular proteins, surface proteins, secretory proteins, exosomes, metabolites or lipids, or any combination thereof. (Item 49) The method according to item 10, wherein the capture probe is located within the polymer matrix. (Item 50) The method according to item 10, wherein the capture probe is located on the surface of the polymer matrix. (Item 51) The method according to item 47, wherein the analyte is released from the cells upon interaction with the reagent. (Item 52) The method according to item 51, wherein the reagent is an oxidizing agent or a reducing agent. (Item 53) The method according to item 51, wherein the reagent is an organic molecule or an inorganic molecule. (Item 54) The method according to item 53, wherein the organic or inorganic molecule is a pharmaceutical compound or a surfactant. (Item 55) The method according to item 51, wherein the reagent is a protein. (Item 56) The method according to item 51, wherein the reagent is a DNA aptamer. (Item 57) The method according to item 51, wherein the reagent is a bead containing a biomolecule. (Item 58) The method according to item 51, wherein the reagent is a biological species. (Item 59) The method according to item 58, wherein the biological species is a virus or a cell. (Item 60) The method according to item 47, wherein the analyte is released from the cells upon exposure to an energy source. (Item 61) The method according to item 60, wherein the energy source is UV light for dissolving cells. (Item 62) The method according to item 60, wherein the energy source is visible light for dissolving cells. (Item 63) The method according to item 61, wherein the UV light is used to activate a photoactivatable surfactant and dissolve the cells. (Item 64) The method according to item 62, wherein the visible light is used to activate a photoactivatable surfactant and dissolve the cells. (Item 65) The method according to item 1, further comprising the step of identifying the analyte or its constituent components. (Item 66) The method according to item 65, wherein the analyte is nucleic acid, amino acid, intracellular protein, surface protein, secretory protein, exosome, metabolite or lipid, or any combination thereof. (Item 67) The method according to item 65, wherein the analyte is a nucleic acid molecule, and the identifying step comprises sequencing of the nucleic acid molecule or its components. (Item 68) The method according to item 1, further comprising the step of measuring the quality of the analyte or its constituent components. (Item 69) The method according to item 68, wherein the quality is the shape or size of the analyte or its constituent components. (Item 70) The method according to item 1, further comprising the step of performing one or more functional assays on the analyte or its components. (Item 71) The method according to item 70, wherein one or more functional assays evaluate cell viability, cell morphology, cell secretion, cell response, cell-cell interactions, or any combination thereof. (Item 72) If the one or more functional assays are colorimetric assays or fluorescence assays, The method described in claim 70. (Item 73) The method according to item 70, wherein the one or more functional assays are performed using bright-field phase-contrast imaging or fluorescence imaging of the analytes. (Item 74) The method according to item 1, further comprising the step of performing one or more omics assays to characterize and quantify the analyte or its components. (Item 75) The method according to item 74, wherein the one or more omics assays are a proteomics assay, a transcriptomics assay, a genomics assay, or an epigenomics assay, or any combination thereof. (Item 76) The method according to item 74, wherein the one or more omics assays are multi-omics assays. (Item 77) A method for processing analytes, (a)(i) providing a fluid device comprising the analyte and (ii) one or more polymer precursors, and (b) A step of configuring a digital micromirror device to direct units of energy generated from an energy source to separate areas of the fluid device to generate a polymer matrix from the one or more polymer precursors within the fluid device, wherein the polymer matrix includes the analyte. Methods that include... (Item 78) The method according to item 77, wherein the fluid device includes a flow channel. (Item 79) The method according to item 77, wherein the fluid device includes an open configuration. (Item 80) The method according to item 77, wherein the fluid device includes one or more distinct locations, and the one or more distinct locations are not in fluid communication with another distinct location. (Item 81) The method according to item 80, wherein the one or more distinct locations are one or more well plates. (Item 82) The method according to item 80, wherein one or more of the aforementioned locations are open at the top. (Item 83) The method according to item 80, wherein the one or more separate locations contain the analyte. (Item 84) The method according to item 77, wherein the fluid device further comprises one or more monomers. (Item 85) The method according to item 77, wherein the fluid device further includes a spatial energy regulating element. (Item 86) The method according to item 77, wherein the fluid device includes a surface on which a capture probe is immobilized, and the capture probe is coupled to the analyte to immobilize the analyte on the surface. (Item 87) The polymer matrix is ​​generated adjacent to or surrounding the analyte. The method described in item 77 for immobilizing the analyte. (Item 88) The method according to item 77, wherein the polymer matrix forms a hydrogel. (Item 89) The method according to item 86, wherein the capture probe comprises one or more functional groups that can interact with the analyte. (Item 90) The method according to item 89, wherein the one or more functional groups include a DNA sequence complementary to the target DNA or RNA. (Item 91) The method according to item 77, wherein the separate area is adjacent to or surrounds the analyte. (Item 92) The method according to item 77, wherein the size of the separate area is adjustable according to the size of the analyte, the shape of the analyte, or other properties of the analyte. (Item 93) The method according to item 77, wherein the aforementioned separate areas are optically identified. (Item 94) The method according to item 77, wherein the detector is configured to detect the location of the analyte within the fluid device. (Item 95) The method according to item 94, wherein the detector configured to detect the location of the analyte within the fluid device is a microscope objective lens for imaging the fluid device. (Item 96) The method according to item 95, wherein an algorithm is used to determine the location of the analyte based on the imaging. (Item 97) The method according to item 95, wherein the imaging is bright-field imaging, phase-contrast imaging, or fluorescence imaging, or any combination thereof. (Item 98) The method according to item 96, wherein the algorithm is a supervised learning algorithm, a self-supervised learning algorithm, or an unsupervised learning algorithm. (Item 99) The method according to item 95, wherein the objective lens is coupled to the energy source which is in optical communication with the fluid device. (Item 100) The method according to item 95, wherein the objective lens directs the digital micromirror device to direct the units of energy to the fluid device. (Item 101) The method according to item 77, further comprising the step of introducing one or more reagents into the polymer matrix that reacts with the analyte. (Item 102) The method according to item 101, wherein one or more of the reagents flow through a membrane. (Item 103) The method according to item 102, wherein the film is semipermeable. (Item 104) The method according to item 102, wherein the membrane includes pores. (Item 105) The method according to item 104, wherein the pore size is less than 10 μm. (Item 106) The method according to item 101, wherein the one or more reagents are an enzyme, an oligonucleotide, a primer, or any combination thereof. (Item 107) The method according to item 101, wherein one or more of the reagents are dissolving reagents. (Item 108) The method according to item 101, wherein one or more of the reagents are nucleic acid denaturing reagents. (Item 109) The method according to item 101, wherein one or more reagents degrade the polymer matrix. (Item 110) The method according to item 77, wherein the analyte is a component of a cell. (Item 111) The method according to item 110, wherein the analyte is a small molecule composed of nucleic acids, amino acids, intracellular proteins, surface proteins, secretory proteins, exosomes, metabolites or lipids, or any combination thereof. (Item 112) The method according to item 86, wherein the capture probe is located within the polymer matrix. (Item 113) The method according to item 86, wherein the capture probe is located on the surface of the polymer matrix. (Item 114) The method according to item 110, wherein the analyte is released from the cells upon interaction with the reagent. (Item 115) The method according to item 114, wherein the reagent is an oxidizing agent or a reducing agent. (Item 116) The method according to item 114, wherein the reagent is an organic molecule or an inorganic molecule. (Item 117) The method according to item 116, wherein the organic or inorganic molecule is a pharmaceutical compound or a surfactant. (Item 118) The method according to item 114, wherein the reagent is a protein. (Item 119) The method according to item 114, wherein the reagent is a DNA aptamer. (Item 120) The method according to item 114, wherein the reagent is a bead containing a biomolecule. (Item 121) The method according to item 114, wherein the reagent is a biological species. (Item 122) The method according to item 114, wherein the biological species is a virus or a cell. (Item 123) The method according to item 110, wherein the analyte is released from the cells upon exposure to an energy source. (Item 124) The method according to item 123, wherein the energy source is UV light for dissolving cells. (Item 125) The method according to item 123, wherein the energy source is visible light for dissolving cells. (Item 126) The method according to item 124, wherein the UV light is used to activate a photoactivatable surfactant and dissolve the cells. (Item 127) The method according to item 125, wherein the visible light is used to activate a photoactivatable surfactant and dissolve the cells. (Item 128) The method according to item 77, further comprising the step of identifying the analyte or its constituent components. (Item 129) The method according to item 128, wherein the analyte is a nucleic acid molecule, and the identifying step comprises sequencing of the nucleic acid molecule or a derivative thereof. (Item 130) The method according to item 77, further comprising the step of measuring the quality of the analyte or its components. (Item 131) The method according to item 130, wherein the quality is the shape or size of the analyte or its constituent components. (Item 132) The method according to item 77, further comprising the step of performing one or more functional assays on the analyte or its components. (Item 133) The method according to item 132, wherein one or more functional assays evaluate cell viability, cell morphology, cell secretion, cell response, cell-cell interactions, or any combination thereof. (Item 134) The method according to item 132, wherein the one or more functional assays are colorimetric assays or fluorescence assays. (Item 135) The method according to item 132, wherein the one or more functional assays are performed using bright-field phase-contrast imaging or fluorescence imaging of the analytes. (Item 136) The method according to item 77, further comprising the step of performing one or more omics assays to characterize and quantify the analyte or its components. (Item 137) The method according to item 136, wherein the one or more omics assays are a proteomics assay, a transcriptomics assay, a genomics assay, or an epigenomics assay, or any combination thereof. (Item 138) The method according to item 136, wherein the one or more omics assays are multi-omics assays. (Item 139) A method for processing analytes, (a)(i) providing a fluid device comprising the analyte and (ii) one or more polymer precursors, and (b) A step of generating a polymer matrix from the one or more polymer precursors using the one or more polymer precursors in the fluid device, wherein the polymer matrix includes the analyte. Includes, (b) is performed in the absence of a physical photomask. method. (Item 140) The method according to item 139, wherein the fluid device includes a flow channel. (Item 141) The method according to item 139, wherein the fluid device includes an open configuration. (Item 142) The method according to item 139, wherein the fluid device includes one or more distinct locations, and the one or more distinct locations are not in fluid communication with another distinct location. (Item 143) The method according to item 142, wherein the one or more distinct locations are one or more well plates. (Item 144) The method according to item 142, wherein one or more of the aforementioned locations are open at the top. (Item 145) The method according to item 142, wherein the one or more distinct locations contain the analyte. (Item 146) The method according to item 139, wherein the fluid device further comprises one or more monomers. (Item 147) The method according to item 139, wherein the fluid device further includes a spatial energy regulating element. (Item 148) The method according to item 139, wherein the fluid device includes a surface on which a capture probe is immobilized, and the capture probe is coupled to the analyte to immobilize the analyte on the surface. (Item 149) The method according to item 139, wherein the polymer matrix is ​​generated adjacent to or surrounding the analyte to immobilize the analyte. (Item 150) The method according to item 139, wherein the polymer matrix forms a hydrogel. (Item 151) The method according to item 148, wherein the capture probe comprises one or more functional groups that can interact with the analyte. (Item 152) The method according to item 151, wherein the one or more functional groups include a DNA sequence complementary to the target DNA or RNA. (Item 153) (b) The method according to item 139, wherein the method comprises exposing one or more polymer precursors to an energy source. (Item 154) The method according to item 153, wherein the energy source is a photogenerating device. (Item 155) The method according to item 154, wherein the light generation device generates light in the range of 350 nm to 800 nm. (Item 156) The method according to item 154, wherein the light generation device generates light in the range of 350 nm to 600 nm. (Item 157) The method according to item 154, wherein the light generation device generates light in the range of 350 nm to 450 nm. (Item 158) The method according to item 154, wherein the photo-generating device generates UV light. (Item 159) (b) The method described in item 139, wherein the method is performed using a spatial light modulator (SLM). (Item 160) The method according to item 159, wherein the SLM is a digital micromirror device (DMD). (Item 161) The method according to item 159, wherein the SLM is a laser beam guided using a galvanometer. (Item 162) The method described in item 159, wherein the SLM is liquid crystal based. (Item 163) The method according to item 139, wherein the polymer matrix is ​​generated in a separate area of ​​the fluid device. (Item 164) The method according to item 163, wherein the separate area is adjacent to or surrounds the analyte. (Item 165) The method according to item 163, wherein the area of ​​the separate area is less than the area of ​​the fluid device. (Item 166) The method according to item 163, wherein the analyte is captured within the separate area. (Item 167) The method according to item 163, wherein the size and shape of the separate areas are adjustable according to the size of the analyte, the shape of the analyte, or other properties of the analyte. (Item 168) The method according to item 167, wherein an algorithm is used to determine the shape and size of the separate areas. (Item 169) The method according to item 168, wherein the algorithm is a supervised learning algorithm, a self-supervised learning algorithm, or an unsupervised learning algorithm. (Item 170) The method according to item 163, wherein the aforementioned separate areas are optically identified. (Item 171) The method according to item 163, wherein the detector is configured to detect the location of the analyte within the fluid device. (Item 172) The method according to item 163, wherein the detector configured to detect the location of the analyte within the fluid device is a microscope objective lens for imaging the fluid device. (Item 173) The method according to item 172, wherein an algorithm is used to determine the location of the analyte based on the imaging. (Item 174) The method according to item 172, wherein the imaging is bright-field imaging, phase-contrast imaging, or fluorescence imaging, or any combination thereof. (Item 175) The method according to item 173, wherein the algorithm is a supervised learning algorithm, a self-supervised learning algorithm, or an unsupervised learning algorithm. (Item 176) The method according to item 172, wherein the objective lens is coupled to an energy source that radiates energy to the separate area in the fluid device. (Item 177) The method according to item 139, further comprising the step of introducing one or more reagents into the polymer matrix that reacts with the analyte. (Item 178) The method according to item 177, wherein one or more of the reagents flow through a membrane. (Item 179) The method according to item 178, wherein the film is semipermeable. (Item 180) The method according to item 178, wherein the membrane includes pores. (Item 181) The method according to item 179, wherein the pore size is less than 10 μm. (Item 182) The method according to item 177, wherein the one or more reagents are an enzyme, a drug molecule, an oligonucleotide, a primer, or any combination thereof. (Item 183) The method according to item 177, wherein one or more of the reagents are dissolving reagents. (Item 184) The method according to item 177, wherein one or more of the reagents are nucleic acid denaturing reagents. (Item 185) The method according to item 177, wherein one or more reagents degrade the polymer matrix. (Item 186) The method according to item 139, wherein the analyte is a component of a cell. (Item 187) The method according to item 186, wherein the analyte is a small molecule composed of nucleic acids, amino acids, intracellular proteins, surface proteins, secretory proteins, exosomes, metabolites or lipids, or any combination thereof. (Item 188) The method according to item 148, wherein the capture probe is located within the polymer matrix. (Item 189) The method according to item 148, wherein the capture probe is located on the surface of the polymer matrix. (Item 190) The method according to item 186, wherein the analyte is released from the cells upon interaction with the reagent. (Item 191) The method according to item 190, wherein the reagent is an oxidizing agent or a reducing agent. (Item 192) The method according to item 190, wherein the reagent is an organic molecule or an inorganic molecule. (Item 193) The method according to item 192, wherein the organic or inorganic molecule is a pharmaceutical compound or a surfactant. (Item 194) The method according to item 190, wherein the reagent is a protein. (Item 195) The method according to item 190, wherein the reagent is a DNA aptamer. (Item 196) The method according to item 190, wherein the reagent is a bead containing a biomolecule. (Item 197) The method according to item 190, wherein the reagent is a biological species. (Item 198) The method according to item 197, wherein the biological species is a virus or a cell. (Item 199) The method according to item 186, wherein the analyte is released from the cells upon exposure to an energy source. (Item 200) The method according to item 199, wherein the energy source is UV light for dissolving cells. (Item 201) The method according to item 199, wherein the energy source is visible light for dissolving cells. (Item 202) The method according to item 200, wherein UV light is used to activate a photoactivatable surfactant and dissolve the cells. (Item 203) The method according to item 201, wherein the visible light is used to activate a photoactivatable surfactant and dissolve the cells. (Item 204) The method according to item 139, further comprising the step of identifying the analyte or its constituent components. (Item 205) The method according to item 204, wherein the analyte is a nucleic acid molecule, and the identifying step comprises sequencing of the nucleic acid molecule or its components. (Item 206) The method according to item 139, further comprising the step of measuring the quality of the analyte or its constituent components. (Item 207) The method according to item 206, wherein the quality is the shape or size of the analyte or its constituent components. (Item 208) The method according to item 139, further comprising the step of performing one or more functional assays on the analyte or its components. (Item 209) The method according to item 208, wherein one or more functional assays evaluate cell viability, cell morphology, cell secretion, cell response, cell-cell interactions, or any combination thereof. (Item 210) The method according to item 208, wherein the one or more functional assays are colorimetric assays or fluorescence assays. (Item 211) The method according to item 208, wherein the one or more functional assays are performed using bright-field phase-contrast imaging or fluorescence imaging of the analytes. (Item 212) One or more omics assays are performed to characterize the analyte or its components. The method according to item 139, further comprising the steps of measuring and quantifying. (Item 213) The method according to item 212, wherein the one or more omics assays are a proteomics assay, a transcriptomics assay, a genomics assay, or an epigenomics assay, or any combination thereof. (Item 214) The method according to item 212, wherein the one or more omics assays are multi-omics assays. (Item 215) A method for analyzing one or more biological components of a biological sample, (a)(i) a channel having an inlet and an outlet, wherein the channel is bounded by a first wall having a first surface and a second wall having a second surface, and the first wall and the second wall are arranged facing each other across the channel; (ii) a spatial energy regulating element arranged adjacent to the first wall, wherein the spatial energy regulating element is capable of projecting energy having predetermined beam characteristics across the channel; (b) The step of loading a reaction mixture containing a biological sample and one or more polymer precursors into the channel, (c) A step of synthesizing one or more chambers in the channels around each of one or more biological components by projecting energy across the channels such that the projected energy crosslinks the one or more polymer precursors to form polymer matrix walls of chambers, Methods that include... (Item 216) The method according to item 215, wherein (i) the fluid device further includes a detector positioned adjacent to the second wall, the detector being capable of detecting an optical signal from the one or more biological components, (ii) the first wall and the second wall each comprise an optically transparent material, and (iii) the projected energy from the spatial energy modulating element includes light. (Item 217) The method according to item 216, wherein the step of synthesizing the chamber includes positioning the chamber to encapsulate one or more biological components based on the optical signal detected by the detector. (Item 218) The method according to item 217, wherein the predetermined beam feature includes a beam having a cross-section that includes a plurality of ring-shaped shapes. (Item 219) The method according to item 217, wherein the predetermined beam feature includes a beam that generates a plurality of chambers, and a portion of the chambers shares the polymer matrix wall with one or more other chambers of the plurality of chambers. (Item 220) The method according to item 217, wherein the polymer matrix wall of the chamber extends from the first surface to the second surface, forming one or more chambers having interiors, respectively. (Item 221) The method according to item 217, wherein the loading step includes stirring the reaction mixture to reduce aggregation of the biological components. (Item 222) The first surface or the second surface is one or more predetermined biological samples of the biological specimen. The method according to item 217, comprising one or more capture elements for specifically capturing biological components, wherein the loading step comprises incubating the reaction mixture under conditions that allow the one or more capture elements to capture the one or more predetermined biological components. (Item 223) The method according to item 217, further comprising the step of removing the reaction mixture after the synthesis step. (Item 224) The method according to item 223, further comprising the step of loading an analytical assay reagent into the channel for determining one or more characteristics of the biological components enclosed in the chamber, wherein the analytical assay reagent can pass through the polymer matrix wall and generate one or more optical signals exhibiting the one or more characteristics. (Item 225) The polymer matrix wall is decomposable by treatment with a decomposing agent, and the method is (a) A step of identifying one or more of the chambers having the selected biological components based on the one or more optical signals from the analytical assay reagent, (b) A step of loading a second reaction mixture containing a second polymer precursor into the channel, wherein the second polymer precursor can form a second polymer matrix wall that is indegradable by at least one degrading agent, and (c) Step of synthesizing a second chamber for enclosing the identified chamber. The method described in item 224, further including the method described in item 224. (Item 226) The method according to item 225, further comprising the step of disassembling the chamber and thereby isolating the biological components having the selected characteristics. (Item 227) A method for analyzing one or more biological components, (a) (i) A channel bounded by a first wall having a first surface, (ii) a spatial energy modulating element disposed adjacent to the first wall, wherein the spatial energy modulating element can project energy having predetermined beam characteristics into the channel across the first wall, step of providing a fluid device having a spatial energy modulating element. (b) The step of loading a reaction mixture containing a biological sample and one or more polymer precursors into the channel, (c) A step of synthesizing one or more chambers in the channels around each of one or more biological components by projecting energy into the channels such that the projected energy crosslinks the one or more polymer precursors to form polymer matrix walls of chambers, Methods that include... (Item 228) The method according to item 227, wherein (i) the fluid device further includes a detector positioned opposite the first wall from the spatial energy modulating element, the detector being capable of detecting an optical signal from the one or more biological components, (ii) the first wall comprises an optically transparent material, and (iii) the projected energy from the spatial energy modulating element includes light. (Item 229) The step of synthesizing the chamber is to synthesize the chamber to contain one or more biological components based on the optical signal detected by the detector. The method of item 228, including positioning.

[0123] Novel properties of the present invention are described in detail in the appended claims. A better understanding of the properties and advantages of the present invention is obtained by reference to the following detailed description which utilizes the principles of the present invention and shows exemplary embodiments, and the accompanying drawings (also referred to herein as "Figure" and "FIG.") are as follows. [Brief explanation of the drawing]

[0124] [Figure 1] Figure 1 shows a schematic diagram of a portion of a channel arranged in a fluid device according to one embodiment.

[0125] [Figure 2A]Figure 2A shows a portion of the system provided herein, including an energy source, according to one embodiment.

[0126] [Figure 2B] Figure 2B shows a polymer matrix formed around a biological component in a portion of the system provided herein, according to some embodiments.

[0127] [Figure 2C] Figure 2C shows a method, according to some embodiments, for forming a polymer matrix around biological components in a system provided herein.

[0128] [Figure 3] Figure 3 is a flowchart illustrating an embodiment of forming a polymer matrix.

[0129] [Figure 4A] Figure 4A shows a portion of a channel containing a trapping element in a fluid device, according to one embodiment.

[0130] [Figure 4B] Figure 4B illustrates a biological component coupled to a trapping element on the surface of a portion of a channel in a fluid device, according to one embodiment.

[0131] [Figure 4C] Figure 4C illustrates a polymer matrix arranged around biological components in a portion of the channel of a fluid device, according to one embodiment.

[0132] [Figure 5] Figure 5 is a flowchart illustrating an embodiment in which a polymer matrix is ​​formed around biological components coupled to the surface.

[0133] [Figure 6A]Figure 6A shows a portion of another embodiment of a fluid device that includes a sealable opening.

[0134] [Figure 6B] Figure 6B shows a method for trapping biological components in a portion of a fluid device according to one embodiment.

[0135] [Figure 7A] Figure 7A shows a schematic top view of a portion of a fluid device according to one embodiment.

[0136] [Figure 7B] Figure 7B shows a schematic top view of a portion of a fluid device containing a polymer matrix, according to one embodiment.

[0137] [Figure 8] Figure 8 shows a schematic top view of a portion of a fluid device containing multiple different reagents, according to one embodiment.

[0138] [Figure 9A] Figure 9A shows a portion of a spatial energy regulating element and a cylindrical polymer matrix according to one embodiment.

[0139] [Figure 9B] Figure 9B shows a polymer matrix in the shape of a hollow cylinder and a portion of a spatial energy regulating element according to some embodiments.

[0140] [Figure 10] Figure 10 shows a micrograph of a polymer matrix compartment containing one or more biological components, according to one embodiment.

[0141] [Figure 11A] Figure 11A illustrates an open compartment formed by a multi-step polymer matrix formation process according to one embodiment.

[0142] [Figure 11B] Figure 11B illustrates a closed compartment formed by a multi-step polymer matrix formation process according to one embodiment.

[0143] [Figure 12A] Figure 12A is a schematic diagram of a portion of the surface of a fluid device coated with a repulsive element, according to one embodiment.

[0144] [Figure 12B] Figure 12B shows a micrograph of biological components captured on a surface using a repulsive element, according to one embodiment.

[0145] [Figure 12C] Figure 12C shows a high-magnification micrograph of biological components captured on a surface using a repulsive element, according to one embodiment.

[0146] [Figure 13A] Figure 13A is a schematic diagram of the cell capture and cell lysis steps in an exemplary mRNA 3' gene expression workflow with external sequencing.

[0147] [Figure 13B] Figure 13B is a schematic diagram illustrating the generation of a cDNA library from template mRNA in an exemplary mRNA 3' gene expression workflow with external sequencing.

[0148] [Figure 13C] Figure 13C is a schematic diagram of sequence library preparation in an exemplary mRNA 3' gene expression workflow involving external sequencing.

[0149] [Figure 14A] Figures 14A and 14B show schematic diagrams of exemplary mRNA 3' gene expression workflows with in situ sequencing. [Figure 14B]Figures 14A and 14B show schematic diagrams of exemplary mRNA 3' gene expression workflows with in situ sequencing.

[0150] [Figure 15] Figure 15 shows a computer system programmed to carry out the methods provided herein, or otherwise configured to do so.

[0151] [Figure 16A] Figure 16A illustrates an example of forming a polymer matrix using an energy-modulating element, according to one embodiment.

[0152] [Figure 16B] Figure 16B shows a top view of an example of a polymer matrix formed using an energy-modulating element according to one embodiment.

[0153] [Figure 17A] Figures 17A–17E illustrate exemplary steps of forming a polymer matrix using a movable energy modulating element, according to some embodiments. [Figure 17B] Figures 17A–17E illustrate exemplary steps of forming a polymer matrix using a movable energy modulating element, according to some embodiments. [Figure 17C] Figures 17A–17E illustrate exemplary steps of forming a polymer matrix using a movable energy modulating element, according to some embodiments. [Figure 17D] Figures 17A–17E illustrate exemplary steps of forming a polymer matrix using a movable energy modulating element, according to some embodiments. [Figure 17E] Figures 17A–17E illustrate exemplary steps of forming a polymer matrix using a movable energy modulating element, according to some embodiments.

[0154] [Figure 18]Figure 18 shows a schematic diagram of a whole-genome workflow in some embodiments.

[0155] [Figure 19] Figure 19 shows a schematic diagram of a multi-omics workflow in some embodiments.

[0156] [Figure 20] Figure 20 shows the polymer matrix inside the well plate format according to some embodiments.

[0157] [Figure 21] Figure 21 shows the polymer matrix inside a 10 mm sized well according to one embodiment.

[0158] [Figure 22] Figure 22 shows a fluid device having a porous membrane on top of a polymer matrix, according to some embodiments.

[0159] [Figure 23] Figure 23 shows a well plate according to one embodiment, which has a hydrogel trap inside the well and a porous membrane on top of the polymer matrix.

[0160] [Figure 24A] Figure 24A shows a microscope equipped with a DMD and an integrated UV irradiation LED, according to one embodiment.

[0161] [Figure 24B] Figure 24B shows a schematic diagram of a DMD in which a virtual mask image is projected onto a fluid channel filled with a polymer precursor that generates a polymer matrix, according to one embodiment.

[0162] [Figure 24C] Figure 24C shows various virtual mask images projected using a DMD according to some embodiments, and the corresponding polymer matrices generated inside the fluid device.

[0163] [Figure 25A] Figure 25A shows a hydrogel structure in which individual cells are encapsulated in a circular hydrogel matrix, according to one embodiment.

[0164] [Figure 25B] Figure 25B shows, in some embodiments, cells identified using bright-field imaging with a 4x microscope objective lens, the resulting Voronoi mask calculated based on the cell location, and the hydrogel structure surrounding the cells generated using the Voronoi mask.

[0165] [Figure 26A] Figure 26A illustrates selective retention of fluorescent cells in a mixture of fluorescent calcein AM-stained and non-fluorescent cells according to some embodiments.

[0166] [Figure 26B] Figure 26B illustrates selective retention of target cells and removal of unwanted cells according to some embodiments.

[0167] [Figure 27A] Figure 27A shows the hydrogel matrix inside a fluid channel with a single cell inside each chamber or CellCage® structure, according to some embodiments.

[0168] [Figure 27B] Figure 27B is a spatial plot showing the locations of all DNA clusters having sequences aligned with mRNA molecules derived from Jurcut cells, according to some embodiments.

[0169] [Figure 27C] Figure 27C shows, using high-magnification images, a highlighted hydrogel matrix with cells inside, and a corresponding spatial plot of DNA sequences mapping to human mRNA, according to one embodiment.

[0170] [Figure 27D] Figure 27D shows, using high-magnification images, a highlighted hydrogel matrix with cells inside, and a corresponding spatial plot of DNA sequences mapping to human mRNA, according to one embodiment.

[0171] [Figure 28A] Figure 28A is a scatter plot showing the number of reads that map specifically to the mouse and human genomes within each hydrogel matrix in some embodiments.

[0172] [Figure 28B] Figure 28B shows spatial plots of DNA sequences mapped to human and mouse mRNA in some embodiments.

[0173] [Figure 29A] Figure 29A shows an oligo sequence, barcode, and antibody label adjacent to a poly(A) tail in some embodiments.

[0174] [Figure 29B] Figure 29B shows the relative expression of various surface proteins detected after sequencing of antibody barcode oligos captured after lysing cells inside a hydrogel matrix, according to some embodiments.

[0175] [Figure 29C] Figure 29C shows a spatial plot of representative cells and barcode sequences inside a cell cage according to some embodiments, where the colors are encoded by the identity of the barcode sequences and illustrate the abundance of various antibodies inside each hydrogel matrix after cell lysis.

[0176] [Figure 30A] Figure 30A shows the fluorescence signal emitted from streptavidin beads that captured IgG molecules secreted from CHO DP-12 cells, according to one embodiment.

[0177] [Figure 30B] Figure 30B shows the fluorescence signal emitted from streptavidin beads that captured an IgG molecule secreted from a single CHO DP-12 cell trapped in a hydrogel matrix, according to one embodiment.

[0178] [Figure 30C] Figure 30C shows a spatial plot of RNA sequences mapping to CHO DP-12 cells trapped in the hydrogel matrix in Figure 30B, according to some embodiments.

[0179] [Figure 31A] Figure 31A shows a schematic diagram of a microfluidic device containing cytokine capture beads and mRNA capture oligos according to some embodiments.

[0180] [Figure 31B] Figure 31B shows gene expression analysis of unstimulated and stimulated Jurcut cells according to some embodiments.

[0181] [Figure 31C] Figure 31C shows fluorescence signals from two hydrogel matrices, each containing a single IL-2-secreting cell, according to one embodiment.

[0182] [Figure 32] Figure 32 shows images of the hydrogel matrix used to culture CHO cells at 0, 18, 42, and 46 hours in some embodiments. [Modes for carrying out the invention]

[0183] This patent or application file includes at least one drawing drawn in color. Copies of this patent or publication of this patent application, accompanied by the color drawing, are available from the Office upon request and payment of the necessary fees.

[0184] Detailed explanation introduction Biological samples can be analyzed at the levels of cellular physical properties, proteome, transcriptome, genome, epigenome, methylome, secretome, or metabolome, either as single cells or in populations of cells. For example, cell-derived genetic material can be analyzed separately from or in combination with genetic material from other cells. Analyzing the components of a sample individually can produce data with higher resolution and less noise or cross-contamination, but this approach can be costly and time-consuming. On the other hand, analyzing one or more samples in bulk can be cost- and time-efficient, but can produce undesirable or less desirable output (e.g., due to heterogeneity). For example, a particular data point may not be traceable back to its source in the sample. Spatial relationships between sample components can also be lost. For example, two components of a sample may be adjacent to each other in their biologically related state, but after pooling multiple samples or components of samples, spatial information between the two components may be lost.

[0185] To avoid the generation of heterogeneous samples, individual components of a sample can be compartmentalized. This may allow for simultaneous or substantially simultaneous processing of individual components of the sample while preserving intact spatial information and reducing the number of processing steps. In addition, by preventing cross-contamination and loss of material during the processing of foreign samples, the generated data may be of higher quality (e.g., the data may have a higher signal-to-noise ratio where appropriate). Some methods may preserve spatial information by barcoding individual compartments or groups of compartments in the sample. These methods may combine barcoded components and may not require multiple assays on the same component within a compartment or analysis chamber. Some of these methods may require a nucleotide amplification step, which may introduce bias into the generated data.

[0186] To compartmentalize individual components of a biological sample, a polymer matrix (e.g., a hydrogel matrix) can be formed adjacent to or around at least a portion of the individual components in a fluid device. The hydrogel matrix may be selectively generated after the system detects a component, surrounding it, or it may be generated in the fluid device according to a predetermined pattern. The hydrogel matrix may allow reagents and small entities to pass through while holding the individual components of the biological sample in place. One or more individual components may be localized (e.g., encapsulated) within the fluid device, and the localized components may be exposed to one or more reagents and / or washing solutions during and / or between analyses, so that multiple assays can be performed within the compartment (e.g., simultaneously, substantially simultaneously, sequentially, etc.).

[0187] Different assays may be performed at different locations in the fluid device, for example, to test the effects of different processing conditions. In addition, since the components are generally not mixed and combined, low concentrations of components (e.g., due to dilution) can be prevented. For example, when analyzing genomic material, the amplification step can be avoided due to the preservation of genomic material in each compartment. Interactions between components can also be studied by having two or more components in a compartment. The polymer matrix may be degradable "on demand," enabling controlled localization and release mechanisms. The solutions provided herein can preserve spatial information of components and generate data at the cellular, proteome, transcriptome, or genomic level. Since spatial information is preserved, the data can be associated with (e.g., concatenated) phenotypic data. Furthermore, the solutions provided herein can preserve spatial information of components and concatenate data (e.g., phenotypic data) at the cellular, proteome, transcriptome, or genomic level.

[0188] While various embodiments of the present invention have been shown and described herein, it will be obvious to those skilled in the art that such embodiments are provided only as examples. A number of variations, modifications, and substitutions can be observed without departing from the present invention. It should be understood that various alternatives to the embodiments of the present invention described herein may be used.

[0189] Whenever the term “at least” precedes the first number in a set of two or more numbers, the term “at least” applies to each of the numbers in that set. For example, at least 1, 2, or 3 is equal to at least 1, at least 2, or at least 3.

[0190] Whenever the term “less than” follows the first number in a set of two or more numbers, the term “less than” applies to each of the numbers in that set. For example, less than 3, 2, or 1 is equal to less than 3, less than 2, or less than 1.

[0191] The terms “coupled,” “connected,” and “communicating with,” as used herein, generally refer to any form of interaction between two or more entities, including mechanical, electrical, magnetic, electromagnetic, fluid, biological, and thermal interactions. Two components may be coupled to one another even if they do not directly contact each other.

[0192] The terms “polypeptide” and “peptide,” when used interchangeably herein, generally refer to polymers of amino acids in which amino acids can be linked to other amino acids by peptide bonds. In some examples, polypeptides are proteins. Amino acids may be naturally occurring amino acids or amino acids that do not exist naturally (e.g., amino acid analogs). Polypeptides may be linear or branched. Polypeptides may contain modified amino acids. Polypeptides may be interrupted by non-amino acids. Polypeptides may exist as single-chain or associated chains. Polypeptides may contain multiple amino acids. Polypeptides may have secondary and tertiary structures (e.g., polypeptides may be proteins). In some examples, polypeptides may contain at least 2, 3, 4, 5, 6, 7, 8, 9, 10, 20, 30, 40, 50, 100, 1,000, 10,000, or more amino acids. Polypeptides may be fragments of larger polymers. In some examples, polypeptides may be fragments of larger polypeptides, e.g., fragments of proteins.

[0193] As used herein, the term "amino acid" generally refers to naturally occurring amino acids or amino acids that do not exist naturally (e.g., amino acid analogs). Amino acids that do not exist naturally may be engineered amino acids or synthetic amino acids.

[0194] As used herein, the term “sample” generally refers to a chemical or biological sample containing biological components. Biological components may include cells, nucleic acids, microbiomes, proteins, cell combinations, metabolites, combinations thereof, or any other suitable components of a biological sample. For example, a sample may be a biological sample containing one or more cells. In another example, a sample may be a biological sample containing one or more polypeptides. A biological sample may be obtained from (e.g., extracted or isolated) blood (e.g., whole blood), plasma, serum, urine, saliva, mucosal exudate, sputum, feces, and tears, or may include such samples. A biological sample may be a fluid or tissue sample (e.g., a skin sample). In some examples, a sample may be derived from a homogenized tissue sample (e.g., brain homogenate, liver homogenate, or kidney homogenate). In certain embodiments, a sample may contain a specific type of cell (e.g., nerve cells, muscle cells, liver cells, or kidney cells). The sample may contain or be obtained from diseased cells or tissues (e.g., tumor cells or necrotic cells). In some embodiments, the sample may contain or be derived from disease-related contaminants (e.g., plaques, biofilms, tumors, or non-cancerous growths). In certain embodiments, the sample may contain or be obtained from cell-free fluids, such as whole blood, saliva, or urine. In various embodiments, the sample may contain circulating tumor cells. In some cases, the sample may contain or be an environmental sample (e.g., soil, waste, or ambient air), an industrial sample (e.g., a sample from any industrial process), or a food sample (e.g., dairy products, vegetable products, or meat products). The sample may be processed before being loaded into a microfluidic device. For example, the sample may be processed to purify a particular cell type or polypeptide and / or to include reagents.

[0195] As used herein, the term “polymer matrix” generally refers to a phase material (e.g., a continuous phase material) containing at least one polymer. In some embodiments, the polymer matrix refers to at least one polymer and the void spaces not occupied by the polymer. The polymer matrix may consist of one or more polymers. The polymer matrix may contain linear, branched, and crosslinked polymer units. The polymer matrix may also contain non-polymer species inserted into the void spaces not occupied by polymer chains. The inserted species may be solid, liquid, or gaseous. For example, the term “polymer matrix” may encompass dry hydrogels, hydrated hydrogels, and hydrogels containing glass fibers. The polymer matrix may also contain polymer precursors, which generally refer to one or more molecules that, upon activation, can cause or initiate polymer reactions. Polymer precursors can be activated by electrochemical energy, photochemical energy, photons, magnetic energy, or any other suitable energy. As used herein, the term “polymer precursor” includes monomers (which polymerize to produce a polymer matrix) and crosslinkable compounds, which may include photoinitiators, other compounds necessary or useful for generating the polymer matrix, and so on.

[0196] As used herein, the term “physical photomask” generally refers to a physical structure having multiple openings or pores through which light can be projected. A physical photomask can be used to polymerize a polymer precursor solution and to form a three-dimensional structure corresponding to a pattern on the photomask, thereby creating the hydrogel matrices described herein. The physical photomask can be patterned in a specific layout or geometric pattern. The physical photomask may be bonded to the upper surface of a flow cell.

[0197] In some embodiments, as used herein, the term “local parameter” means a parameter (such as pH) value in or immediately adjacent to a chamber formed by a polymer matrix wall.

[0198] As used herein, the term “on demand” means that operations can be directed to individual, separate, selected locations (e.g., spatial locations of polymer precursor solutions or selected polymer matrix chambers). Such selection may be based on manual observation of optical signals or data collected by a detector, or on computer algorithms that operate on optical signals or data collected by a detector. Manual observation of optical signals or data collected by a detector may include real-time detection or detection at some time before adjusting the energy units for polymerizing the polymer precursor or before decomposing the chambers. For example, a subset of chambers (all formed by photodegradable polymer matrix walls) may be pre-selected to release and remove their contents based on location information and the values ​​of optical signals from analytical assays performed in the chambers. Pre-selected chambers may be photodegraded by selectively projecting rays with appropriate wavelength characteristics to decompose the polymer matrix walls of the pre-selected chambers (e.g., by a spatial energy modulating element). In another example, multiple chambers may be observed in real time (e.g., via a fluorescence microscope) for the detection of the target analyte, and one or more of the multiple chambers may be selected in real time for decomposition when detecting the target analyte.

[0199] Where used herein, the terms “microfluidics” and “nanofluidics” in reference to devices are used interchangeably herein and refer to an integrated system for capturing, moving, mixing, dispersing, or analyzing small volumes of fluid, including, but not limited to, a sample (which may then contain or constitute the cell or molecular analyte of interest), reagents, dilutants, buffers, etc. Generally, references to “microfluidics” and “nanofluidics” represent different scales in device size and the volume of fluid handled. In some embodiments, the properties of a microfluidic device have cross-sectional dimensions of less than several hundred square micrometers, and have passages or channels with capillary dimensions, e.g., with a maximum cross-sectional dimension of about 500 μm to about 0.1 μm. In some embodiments, the microfluidic device has a volumetric capacity ranging from 1 μL to several nL, e.g., 10 to 100 nL. The dimensions of the corresponding properties or structures in a nanofluidic device are typically one to three orders of magnitude smaller than the dimensions of the microfluidic device. Those skilled in the art will know the number of dimensions from the context of the particular application to which they relate. In some embodiments, a microfluidic device or nanofluidic device has one or more interconnected and fluid-communicating chambers, ports, and channels, designed to perform one or more analytical reactions or processes, either alone or in cooperation with instruments or equipment that provide supporting functions such as sample introduction, fluid and / or reagent driving means, e.g., positive or negative pressure, acoustic energy, temperature control, detection systems, data acquisition and / or integration systems. In some embodiments, the microfluidic and nanofluidic devices may further include valves, pumps, filters, and special functional coatings on the inner walls to prevent adsorption of sample components or reactants, facilitate the movement of reagents by electroosmosis, etc.Such devices may be manufactured as integrated devices on a solid substrate, which may be glass, plastic, or other solid polymer material, and may have a planar format for ease of sample detection and monitoring and reagent transfer, particularly via optical or electrochemical methods. In some embodiments, such devices are disposable after single use. The fabrication and operation of microfluidic and nanofluidic devices are well known in the art, as exemplified by the following references incorporated by reference: Ramsey, U.S. Patent Nos. 6,001,229; 5,858,195; 6,010,607; and 6,033,546; Soane et al., U.S. Patents Nos. 5,126,022 and 6,054,034; Nelson et al., U.S. Patent No. 6,613,525; Maher et al., U.S. Patent No. 6,399,952; Ricco et al., International Patent Publication No. WO02 / 24322; Bjornson et al., International Patent Publication No. WO99 / 19717; Wilding et al., U.S. Patent Nos. 5,587,128; and 5,498,392; Sia et al, Electrophoresis, 24: 3563-3576 (2003); Unger et al, Science, 288: 113-116 (2000); Enzelberger et al, U.S. Patent No. 6,960,437; Cao, “Nanostructures & Nanomaterials: Synthesis, Properties & Applications” (Imperial College Press, London, 2004); Haeberle et al, LabChip, 7: 1094-1110 (2007); Cheng et al, Biochip Technology (CRC Press, 2001), etc.

[0200] As used herein, the term “analyte” generally refers to a distinct biological or chemical entity measured, detected, and / or identified using the methods and systems described herein. In some embodiments, the analyte may be a biological component described herein.

[0201] System for the analysis of biological components This disclosure provides a system for partitioning or isolating one or more biological components. The system may include a fluid device containing or comprising one or more biological components. The fluid device may contain or comprise one or more polymer precursors. In some cases, the fluid device may include a first surface configured to couple with or accept at least one of the one or more biological components to form a coupled biological component. The system may also include at least one energy source, which is in communication with the fluid device. In some embodiments, the energy source may be in optical communication with the fluid device. In various embodiments, at least one energy source may form a polymer matrix on or adjacent to at least a portion of one or more biological components.

[0202] In some cases, the sample may be introduced into or provided to the system. In certain cases, the sample may contain one or more biological components. The system may be used to separate one or more biological components from each other. In various cases, the biological components may be physically separated. In some cases, the biological components may be in fluid communication with each other. In certain cases, the biological components may be in chemical communication with each other. The system may be used for single-cell analysis. In some embodiments, the system may be used for single-cell analysis at the genome level. For example, the system may be used for genome sequencing. In another example, the system may be used for deoxyribonucleic acid (DNA) sequencing. The system may be used for DNA sequencing of cell-free DNA, whole-genome sequencing, whole-exome sequencing, targeted sequencing, or 16S sequencing. The system may be used to study DNA tags attached to biomolecules of interest. Biomolecules may include proteins, metabolites, etc. In some cases, the DNA may be nuclear DNA or mitochondrial DNA. The system may be used for single-cell or bulk analysis at the transcriptome level. For example, the system may be used for ribonucleic acid (RNA) sequencing. For example, the system may be used for 3' or 5' gene expression analysis, cellular immunorepertory studies, or full-length mRNA analysis. In some embodiments, the system may be used for single-cell analysis at the proteome level. The system may be used for functional assays of biological components. The system may be used to study surface proteins, secreted proteins, or metabolites of biological components. In some cases, the system may be used to measure the quality of biological components. In some cases, the quality measured may be the size or shape of the biological component. In some cases, the system may be used to study epigenomics, DNA methylation, or chromatin accessibility in biological components.The system may be used for other suitable assays, experiments, and processes.

[0203] In certain embodiments, the system may be used for single-cell analysis at the level of indirect intercellular interactions. For example, the effect of one or more molecules produced from a first cell on a second cell can be analyzed using the system provided herein. In various embodiments, the system may be used to analyze direct intercellular interactions. For example, two or more cells (e.g., a first cell and a second cell) can be in physical contact, and the effect of one or more of the first cell on the second cell, or vice versa, can be analyzed using the system disclosed herein. In some embodiments, the system may be used for drug response analysis of biological components. In certain embodiments, the system may be used to analyze the response of biological components to various physiological conditions (e.g., various culture media, temperatures, mechanical stimuli, etc.).

[0204] In some cases, the sample includes a biological sample. The biological sample may contain biological components. In some embodiments, the biological sample is 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, 30, 40, 50, 100, 200, 300, 400, 500, 600, 700, 1,000, 10,000, 10 5 , 10 6 , 10 7 , 10 8 , 10 9 , 10 10 , 10 20 The biological sample may contain 10 or more biological components. The biological sample may contain any number of biological components between any two of the numbers referred to herein. In some embodiments, the biological sample may contain 10 20It may contain more biological components than those listed above. The biological components may include cells. In some embodiments, the cells may include eukaryotic cells, prokaryotic cells, fungal cells, protists, algal cells, plant cells, animal cells (e.g., human cells), or any other suitable cells. The biological components may include cells, viruses, bacteria, nucleic acids (e.g., DNA or RNA), proteins, or combinations thereof. The combinations may include DNA-protein complexes, RNA-protein complexes, or combinations thereof. In certain embodiments, the nucleic acids may include DNA. The DNA may be at least 10 base pairs (bp) long. In some embodiments, the DNA may be at least 10 bp, 20 bp, 30 bp, 40 bp, 50 bp, 60 bp, 70 bp, 80 bp, 90 bp, 100 bp, 200 bp, 300 bp, 400 bp, 500 bp, 600 bp, 700 bp, 800 bp long, or longer than 800 bp.

[0205] Polymer precursors can be formed using any of the hydrogel precursors and crosslinking agents (first and third columns, respectively) in Table 1A. The resulting polymer matrix can be degraded by the degrading agents shown in Table 1A (fourth column). [Table 1A] [Table 1B]

[0206] In certain embodiments, one or more polymer precursors may be added to or included with a biological sample. One or more biological samples and one or more polymer precursors may be introduced into a system (e.g., into a fluid device of the system). One or more biological samples and one or more polymer precursors may be introduced into the fluid device in any order (e.g., in parallel, sequentially, etc.). For example, the biological sample may be introduced before the polymer precursors, the polymer precursors may be introduced before the biological samples, and the biological samples and polymer precursors may be introduced simultaneously (or substantially simultaneously) or in any other preferred manner or order. In some embodiments, the polymer precursors may comprise one or more hydrogel precursors. One or more polymer precursors may be stored and / or introduced into the system separately. In some cases, one or more polymer precursors may be mixed with one or more biological components before introduction into the system. In various cases, one or more polymer precursors may be mixed with one or more biological components after introduction into the system.

[0207] The system may include a fluid device. In some embodiments, the fluid device may include one or more polymer precursors. In other words, one or more polymer precursors may be located within at least a portion of the fluid device (for example, within at least a portion of the channels of the fluid device). In some embodiments, the fluid device may include one or more channels or chambers. In some embodiments, the fluid device may include at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, 30, 40, 50, 100, 200, 300, 400, 500, 600, 700, 1,000, 10,000 channels or chambers, or any number between any two of the numbers referred to herein. In some embodiments, the fluid device includes more than 10,000 channels or chambers. As described herein, the fluid device may include one or more channels. The fluid device may also, or alternatively, include one or more chambers. The terms channel and chamber may be used interchangeably in this disclosure unless otherwise indicated. For example, a channel or chamber in a fluid device may include a first surface, a second surface, or more surfaces.

[0208] The channel or chamber of the fluid device (sometimes also referred to as a “flow chamber” or “reaction chamber” as opposed to a chamber formed from polymer matrix walls within the channel) may accept or be configured to accept a biological sample. Figure 1 shows a schematic diagram of a portion of a channel 100 that may be located in at least a portion of the fluid device of a system provided herein. The fluid device may include the channel 100. The channel 100 may include a first surface 101. Furthermore, the channel 100 may include a second surface 102. In some embodiments, the first surface 101 and the second surface 102 are positioned, installed, or located facing each other (e.g., as shown in Figure 1). In some embodiments, the first and second surfaces are substantially parallel, and as a result, the perpendicular distance between them is substantially the same across the entire channel in which the chamber is formed, for example. In some embodiments, the perpendicular distance between the first and second surfaces depends in part on the characteristics and size of the biological components being analyzed. In some embodiments, for example, embodiments adapted for the analysis of mammalian cells, the vertical distance between the first and second surfaces may be in the range of 10 μm to 500 μm, or 50 μm to 250 μm. In some embodiments, the vertical distance between the first and second surfaces may be in the range of twice the average size of the biological components being analyzed to five times the average size of the biological components being analyzed. In some embodiments, the vertical distance between the first and second surfaces may be in the range of twice the average size of the largest biological component in the biological sample to five times the average size of the largest biological component in the biological sample. In some embodiments, the first surface 101 may be the bottom surface. In certain embodiments, the second surface 102 may be the top surface. The terms “bottom” and “top” are used herein for convenience, not as limiting, with reference to the drawings. Channel 100 may accept a biological sample containing one or more biological components 50, 51. Channel 100 may accept one or more polymer precursors. As illustrated in Figure 1, the biological components 50, 51 may include cells.However, as discussed herein, the biological components may include tissues, proteins, nucleic acids, and the like. In some embodiments, the first surface 101, the second surface 102, or both surfaces may be capable of coupling to or accepting at least one of one or more biological components 50, 51, or configured to couple to or accept it. In some cases, the first surface 101 may be capable of coupling to or accepting a biological component (e.g., biological components 50, 51), or configured to couple to or accept it. In certain particular cases, the second surface 102 may be capable of coupling to or accepting a biological component (e.g., biological components 50, 51), or configured to couple to or accept it.

[0209] In certain cases, the channel may have a rectangular, circular, semicircular, elliptical, or other preferred cross-section. Thus, the channel may have a single internal surface. In some cases, the channel may have a triangular, square, rectangular, polygonal, or other cross-section. Thus, the channel may have three or more internal surfaces. One or more internal surfaces may be able to couple to or accept one or more biological components, or may be configured to couple to or accept them.

[0210] In some cases, the first surface 101, the second surface 102, or both surfaces 101, 102 may be functionalized with, for example, a coating (e.g., a surface coating). In some embodiments, the surface coating may be a surface polymer. Some non-limiting examples of surface coatings include capture reagents (e.g., pyridine carboxyaldehyde (PCA)), functional groups for capturing one or more moieties (e.g., chemical moieties), functional groups including acrylamide, agarose, biotin, streptavidin, strep-tag II, linkers, aldehydes, phosphates, silicates, esters, acids, amides, alkynes, azides, aldehyde dithiolanes, or combinations thereof. In various embodiments, the surface coating may contain functional groups for capturing one or more moieties. For example, acrylamide, agarose, etc., may contain such functional groups. In certain embodiments, the surface polymer may contain polyethylene glycol (PEG), thiols, alkenes, alkynes, azides, or combinations thereof. In various embodiments, the surface polymer may include a silane polymer. In some embodiments, the surface polymer may be functionalized with at least one of oligonucleotides, antibodies, cytokines, chemokines, proteins, antibody derivatives, antibody fragments, carbohydrates, toxins, or aptamers.

[0211] In some cases, the first surface 101, the second surface 102, or both surfaces 101, 102 may contain one or more barcodes (e.g., nucleic acid barcodes). In some embodiments, the first surface 101, the second surface 102, or both surfaces 101, 102 may contain 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, 30, 40, 50, 100, 200, 300, 400, 500, 600, 700, 1,000, 10,000, 50,000, 100,000, 250,000, 500,000, 1,000,000, 2,000,000, 5,000,000, 10,000,000, 15,000,000 barcodes, or any number of barcodes between any two numbers mentioned herein. The barcode is about 500 nm 2 ~ about 500 μm 2The area may be covered. In some embodiments, the first surface 101, the second surface 102, or both surfaces 101, 102 may contain a total number of barcodes up to about 10,000,000. The barcodes may be different from each other (for example, each barcode may be unique). In certain embodiments, the first part or subset of the barcode may be different from the second part or subset of the barcode. There may be 2, 3, 4, 5, 10, 15, 20, 25, 50, 75, 100, 1,000, 10,000 parts or subsets of the barcode, or any number of parts or subsets of the barcode between any two numbers referred to herein. In some cases, the barcode (or part / subset of the barcode) may be associated with the location of the barcode on the surface (the coordinates of the location on the channel's surface (e.g., x, y coordinates)). The barcode may be attached to or coupled to the captured biological component. In some embodiments, the barcode may be a unique identifier that distinguishes a biological component from other biological components (e.g., identifying a first biological component relative to a second biological component). In some embodiments, the barcode may include a nucleic acid sequence (e.g., a common sequence) for capturing the biological component, or may be used in amplification. In some embodiments, the barcode may include a unique identifier that includes a unique nucleic acid sequence (e.g., a DNA sequence, an RNA sequence, etc.), a protein tag, an antibody, or an aptamer. In some embodiments, the barcode may include a fluorescent molecule. In some embodiments, the location of the captured biological component may be associated with the unique identifier, for example, to preserve spatial information of the biological component.

[0212] In some embodiments, the fluid device may be a flow cell. For example, the fluid device may be used for sequencing (e.g., DNA or RNA sequencing). In some embodiments, the fluid device may be a microfluidic device. In certain embodiments, the fluid device may be a nanofluidic device.

[0213] The systems disclosed herein may include one or more energy sources. The energy sources may be in communication with the fluid device. In some embodiments, the energy sources may be in optical communication with the fluid device. In some cases, the energy sources can be used to form one or more polymer matrices in the fluid device (e.g., on or adjacent to the surface of a channel or chamber of the fluid device). In some embodiments, the energy sources may include a photogenerating device, a thermogenerating device, an electrochemical reaction generating device, an electrode, or a microwave device. The polymer matrices may be formed in the channels of the fluid device. The energy sources may direct or move energy to a predetermined location in the fluid device. At a predetermined location, the energy may cause (e.g., polymerize) one or more polymer precursors to form a polymer matrix, or may be activated to form one or more polymer precursors.

[0214] In some embodiments, the polymer matrix may include a hydrogel. In some embodiments, the hydrogel may be sufficiently porous to allow the movement or migration of reagents (e.g., enzymes, chemical compounds, small molecules, antibodies, etc.) through the polymer matrix, or may have pores of a suitable size, but the hydrogel does not have to allow the movement or migration of biological components (e.g., DNA, RNA, proteins, cells, etc.) through the polymer matrix. In some embodiments, the pores may have a diameter of 5 nm to 100 nm. In some embodiments, the pores may have a diameter of 5 nm to 10 nm, 10 nm to 20 nm, 20 nm to 30 nm, 30 nm to 40 nm, 50 nm to 60 nm, 60 nm to 70 nm, 70 nm to 80 nm, 80 nm to 90 nm, or 90 nm to 100 nm. In some embodiments, the pores may have a diameter greater than 100 nm. In some embodiments, the pores may have a diameter less than 5 nm. The reagent may include an enzyme or a primer having a size of less than 50 base pairs (bp). The primers may contain single-stranded DNA (ssDNA). In some embodiments, the primers may have a size of 5 bp to 50 bp. In some embodiments, the primers may have a size of 5 bp to 10 bp, 10 bp to 20 bp, 20 bp to 30 bp, 30 bp to 40 bp, or 40 bp to 50 bp. In some embodiments, the primers may have a size greater than 50 bp. In certain particular cases, the primers may have a size less than 5 bp. The reagents may include lysozyme, proteinase K, hexamers (e.g., random hexamers), polymerases, transposases, ligases, catalytic enzymes, deoxyribonucleases, deoxyribonuclease inhibitors, ribonucleases, ribonuclease inhibitors, DNA oligos, deoxynucleotide triphosphates, buffers, surfactants, salts, divalent cations, or any other suitable reagents.

[0215] In some embodiments, one or more reagents flow through the membrane. In some embodiments, the membrane is semipermeable. In some embodiments, the membrane contains pores. In some embodiments, the pores are less than 1 micrometer wide. In some embodiments, the width of the pores is about 0.5 micrometers to about 15 micrometers. In some embodiments, the width of the pores is about 0.5 micrometers to about 1 micrometer, about 0.5 micrometers to about 5 micrometers, about 0.5 micrometers to about 10 micrometers, about 0.5 micrometers to about 15 micrometers, about 1 micrometer to about 5 micrometers, about 1 micrometer to about 10 micrometers, about 1 micrometer to about 15 micrometers, about 5 micrometers to about 10 micrometers, about 5 micrometers to about 15 micrometers, or about 10 micrometers to about 15 micrometers. In some embodiments, the width of the pores is about 0.5 micrometers, about 1 micrometer, about 5 micrometers, about 10 micrometers, or about 15 micrometers. In some embodiments, the pore width is at least about 0.5 micrometers, about 1 micrometer, about 5 micrometers, or about 10 micrometers. In some embodiments, the pore width is at most about 1 micrometer, about 5 micrometers, about 10 micrometers, or about 15 micrometers. In some embodiments, one or more reagents are enzymes, drug molecules, oligonucleotides, primers, or any combination thereof. In some embodiments, one or more reagents are dissolving reagents. In some embodiments, one or more reagents are nucleic acid denaturing reagents. In some embodiments, one or more reagents degrade the polymer matrix.

[0216] Figure 22 shows a fluid device having a porous membrane 2210 on top. The biological component 2202 is encapsulated and / or localized on the surface of the fluid device 2206 using a polymer matrix 2204. Reagents passing through the pores in the membrane 2210 can enter the polymer matrix and react with the biological component 2202. In some embodiments, as shown in Figure 23, the polymer matrix 2304 and the biological component 2302 are seeded in a well plate 2306, and the porous membrane 2310 is on top of the polymer matrix 2304. Reagents passing through the pores in the membrane 2310 can enter the polymer matrix and react with the biological component 2302. Due to the walls of the wells 2308, fluid communication between different wells may not be necessary, so that different reagents can be used in each individual well.

[0217] Figure 2A shows a portion of the system provided herein, including an energy source 203. The embodiment in Figure 2A may include components similar in some respects to the components in Figure 1. For example, the embodiment in Example 2A includes a channel 200 which may be similar to channel 100 in Figure 1. It will be recognized that the illustrated embodiments may have similar characteristics. Accordingly, similar characteristics are designated by similar reference numbers, with the leading digit incrementing to "2". The relevant disclosures shown above with respect to similarly identified characteristics are thus not repeated below in this specification. Also, certain characteristics of the system provided herein and the relevant components shown in Figure 2A may not be indicated or identified by reference numbers which are specifically discussed in the drawings or in the subsequent specification. However, such characteristics may be explicitly the same or substantially the same as those shown in other embodiments and / or described for such embodiments. Accordingly, the relevant descriptions of such characteristics apply equally to the characteristics of the system and relevant components in Figure 2A. Any preferred combination of the properties and variations thereof described for the system and components illustrated in Figure 1 can be used with the system and components of Figure 2A, and vice versa. This pattern of disclosure is equally applicable to further embodiments shown in later drawings and described below in this specification.

[0218] Continuing with Figure 2A, the system channel 200 may include a first surface 201 and a second surface 202. In some embodiments, the energy source 203 may include one or more energy-emitting portions (e.g., energy-emitting portion 205). In some embodiments, the energy source 203 may include one or more non-emitting portions (e.g., non-emitting portion 204). The non-emitting portion 204 may not emit energy, nor may it be configured to emit energy. In some embodiments, the emitting portion 205 may radiate energy to at least one part of the fluid device in the form of electromagnetic waves (e.g., microwaves, light, heat, etc.). In certain embodiments, the emitting portion 205 may radiate energy to the fluid device. In some embodiments, the fluid channel may be coupled to a movable stage. In other embodiments, light may be projected onto at least one part of the fluid channel to generate one or more polymer matrices. The light may be directed to different parts of the fluid channel. In some embodiments, the radiating portion 205 may be coupled to an objective lens (e.g., a microscope objective lens or lens), where the objective lens may move to different parts of the fluid device. The objective lens may provide a shape (e.g., a virtual mask or a physical mask) that allows light to form a pattern on the fluid device to form a polymer matrix similar to or complementary to the pattern. In various embodiments, one or more polymer precursors in the fluid device or mixed with a biological sample may absorb the radiant energy 206. In some embodiments, the radiant energy 206 may be able to form a polymer matrix from one or more polymer precursors, or may be sufficient to form one. For example, a portion of one or more polymer precursors in a channel 200 of the fluid device may be activated by the radiant energy, initiating a polymerization reaction to form a polymer matrix.

[0219] In some embodiments, the energy source may radiate energy to a large portion of the fluid channel or to substantially the entire surface of the fluid channel. A physical mask may be used to block energy radiated to one or more portions of the fluid channel. The energy source (e.g., a light source) may be coupled to the fluid device via an objective lens (e.g., a microscope objective lens or lens). The energy source may be directed to a portion of the fluid channel (e.g., via a movable objective lens). In some cases, the light source, objective lens, and / or fluid channel are movable to allow energy to be radiated to the fluid channel in such a way that it generates a pattern on at least a portion of the surface of the fluid device. The polymer matrix may be formed in a manner similar to or complementary to the energy radiation pattern.

[0220] The polymer precursor may include an activating molecule that can absorb the radiant energy 206 that initiates polymerization of one or more polymer precursors in the fluid device. Non-limiting examples of activating molecules may include photocatalysts, photoactivators, photoacid generators, or photobase generators. In some embodiments, the first polymer matrix 208 and / or the second polymer matrix 209 may be formed on or adjacent to the biological component 50. In certain embodiments, the first polymer matrix 208 and the second polymer matrix 209 may form an analytical chamber or compartment 220 in the fluid device that separates (e.g., physically separates) the biological component 50 from other biological components (e.g., biological components 51, 52, or 53). In other words, the polymer matrix may compartmentalize a channel (e.g., channel 200). In various embodiments, the polymer matrix may partially surround the biological component. For example, the polymer structure surrounding a biological component may form a closed structure (e.g., a hollow cylindrical polymer structure) or a partially open structure (e.g., a crescent-shaped polymer structure). In some embodiments, two or more polymer matrices may be formed adjacent to a biological component to form a compartment that separates the biological component from other biological components. In certain embodiments, the polymer matrix may include or form a wall (e.g., a polymer matrix wall).

[0221] In various embodiments, the polymer matrix comprises a hydrogel. In some embodiments, the polymer matrix wall may be a hydrogel wall. In some embodiments, the hydrogel or hydrogel wall may be polyethylene glycol (PEG)-thiol, PEG-acrylate, acrylamide, N,N'-bis(acryloyl)cystamine (BAC), PEG, polypropylene oxide (PPO), polyacrylic acid, poly(hydroxyethyl methacrylate) (PHEMA), poly(methyl methacrylate) (PMMA), poly(N-isopropylacrylamide) (PNIPAAm), poly(lactic acid) (PLA), poly(lactic acid-co-glycolic acid) (PLGA), polycaprolactone (PCL), poly(vinyl sulfonic acid) (PVSA), poly(L-aspartic acid), poly(L-glutamic acid), polylysine, agar, It may contain agarose, alginate, heparin, alginate sulfate, dextran sulfate, dextran-acrylamide, hyaluronan, pectin, carrageenan, gelatin, chitosan, cellulose, collagen, bisacrylamide, diacrylate, diallylamine, triallylamine, divinyl sulfone, diethylene glycol diallyl ether, ethylene glycol diacrylate, polymethylene glycol diacrylate, poly(ethylene glycol) diacrylate, trimethylopropane trimethacrylate, ethoxylated trimethylol triacrylate, ethoxylated pentaerythritol tetraacrylate, or combinations or mixtures thereof. The hydrogel or hydrogel wall may contain a degradable crosslinking agent (e.g., N,N'-bis(acryloyl)cystamine, chitosan, poly(ε-caprolactone) diacrylate, polylactide diacrylate, polylactide dimethacrylate, poly(lactide-co-glycolide), polycaprolactone molecule, or other suitable degradable crosslinking agents).

[0222] In some embodiments, the surface of the polymer matrix or hydrogel may be functionalized by coupling functional groups to the polymer matrix or hydrogel. Some non-limiting examples of functional groups include capture reagents (e.g., pyridine carboxyaldehyde (PCA)), acrylamide, agarose, biotin, streptavidin, strep-tag II, linkers, aldehydes, phosphates, silicates, esters, acids, amides, aldehyde dithiolanes, PEGs, thiols, alkenes, alkynes, azides, or combinations thereof. In some cases, a functionalized polymer matrix may be used to capture biomolecules inside polymer matrix compartments formed adjacent to (e.g., around or on) biological components. The biomolecules may be produced by the biological components (e.g., cell-derived secretomes). The functionalized surface of the polymer matrix inside the compartment may be used to capture reagents or molecules from outside the compartment. The functionalized surface may increase the surface area that can be covered by reagents, molecular sensors, or any molecule of interest (e.g., antibodies).

[0223] In some embodiments, the compartments surrounding the biological components may include a polygonal base. In various embodiments, the compartments surrounding the biological components may include a circular or elliptical base (see, for example, compartment 220 or compartment 222 in Figure 2C). In certain embodiments, the polymer matrix walls of the compartments may have a thickness (e.g., width) of 1 μm to 250 μm. Polymer matrix walls or compartments are 1μm~5μm, 1μm~10μm, 1μm~20μm, 1μm~30μm, 1μm~40μm, 1μm~50μm, 1μm~100μm, 1μm~150μm, 1μm~250μm, 5μm~10μm, 5μm~20μm, 5μm~30μm, 5μm~40μm, 5μm~50μm, 5μm~100μm, 5μm~150μm, 5μm~250μm, 10μm~20μm, 10μm~30μm, 10μm~40μm, 10μm~50μm, 10μm~100μm, 10μm~150μm, 10μm~250 It may have a thickness of μm, 20μm~30μm, 20μm~40μm, 20μm~50μm, 20μm~100μm, 20μm~150μm, 20μm~250μm, 30μm~40μm, 30μm~50μm, 30μm~100μm, 30μm~150μm, 30μm~250μm, 40μm~50μm, 40μm~100μm, 40μm~150μm, 40μm~250μm, 50μm~100μm, 50μm~150μm, 50μm~250μm, 100μm~150μm, 100μm~250μm, or 150μm~250μm. The polymer matrix walls or compartments may have a thickness of approximately 1 μm, 5 μm, 10 μm, 20 μm, 30 μm, 40 μm, 50 μm, 100 μm, 150 μm, or 250 μm. The polymer matrix walls or compartments may have a thickness of at least 1 μm, 5 μm, 10 μm, 20 μm, 30 μm, 40 μm, 50 μm, 100 μm, 150 μm, 250 μm, or greater. The polymer matrix walls or compartments may have a maximum thickness of 5 μm, 10 μm, 20 μm, 30 μm, 40 μm, 50 μm, 100 μm, 150 μm, or 250 μm.The polymer matrix walls or compartments may have a thickness of less than 1 μm.

[0224] Continuing with Figure 2A, polymer matrices 208, 209, or at least a portion of polymer matrices 208, 209, may be coupled to a first surface 201, a second surface 202, or both surfaces 201, 202. In certain embodiments, the polymer matrix, or at least a portion of the polymer matrix, may be coupled to a third surface, a fourth surface, a fifth surface, etc., where appropriate. In various embodiments, polymer matrices 208, 209 may extend from the first surface 201 to the second surface 202 such that the polymer matrix surrounds or substantially surrounds the biological component 50 (e.g., through at least a portion of the lumen or chamber cavity of the channel 200). In some embodiments, two or more biological components that are physically close together (e.g., biological components 50, 51 in Figure 2C) may be separated (e.g., by stirring or shaking the fluid device). The fluid device may be agitated or shaken by physical motion, the use of sonic pulses, altering the flow in the channel, or any other suitable method of agitation. A polymer matrix may then be formed surrounding (or partially surrounding) the biological components to be separated. Figure 2B shows polymer matrices 208, 209 formed surrounding biological component 50 after separation from biological component 51. Figure 2C illustrates the process of separating two adjacent biological components 50, 51 according to various embodiments; that is, the biological components 50, 51 can be separated by agitating or shaking the fluid device. In some embodiments, the separation of biological components is achieved by fluid pressure, flow pulsation, dielectrophoresis, optothermal flow, or a combination of some of these. In some cases, the separation of biological components is achieved by acoustic vibration. Figure 2C shows a polymer matrix formed to generate a compartment 222 surrounding biological component 50 after the separation of biological components 50, 51.

[0225] Continuing with Figure 2A, in some cases, the energy source 203 may form or produce one or more radiating portions 205 and one or more non-radiating portions 204, or may be configured to form or produce them. The systems disclosed herein may further include a spatial energy modulator that directs energy from the energy source to one or more targeted portions of the fluid device. For example, the spatial energy modulator may be configured to selectively direct energy from the energy source to form a polymer matrix in a separate area of ​​the fluid device. In some embodiments, the separate area is selected based on the location of a biological component. In some embodiments, the area of ​​the separate area is less than the area of ​​the fluid device. In some embodiments, the biological component is trapped within the separate area. In some embodiments, the size and shape of the separate area are adjustable according to the size, shape, or other properties of the biological component. In some embodiments, an algorithm is used to determine the shape and size of the separate area. In some embodiments, the algorithm is a supervised learning algorithm, a self-supervised learning algorithm, or an unsupervised learning algorithm. The spatial energy modulating element may be configured to selectively direct energy, for example, by blocking or preventing energy from being directed to one or more parts of a fluid device other than one or more targeted parts. In some embodiments, the spatial energy modulating element may include a physical mask. In some embodiments, the spatial energy modulating element may include a virtual mask. In some cases, the spatial energy modulating element may be a spatial light modulator (SLM). In some embodiments, the SLM is a digital micromirror device (DMD). In some embodiments, the SLM is a laser beam guided using a galvanometer. In some embodiments, the SLM is liquid crystal based.

[0226] Figure 24A shows a microscope with a DMD and an integrated UV irradiation LED, according to one embodiment. Figure 24B shows a DMD setup, according to one embodiment, projecting a virtual mask image onto a channel filled with a polymer precursor that generates a polymer matrix. Figure 24C shows various virtual mask images projected using the DMD and the corresponding hydrogel structures generated inside a fluid device.

[0227] In certain cases, the spatial energy modulating element may be configured to control one or more electrodes that can selectively deliver energy to one or more targeted portions of a fluid device. Using the electrode concept, spatially regulated energy may be provided to form a hydrogel structure. In some implementations, one or more electrodes can be arranged in predetermined locations within a fluid channel, thus enabling the formation of a hydrogel at these locations. In alternative implementations, the electrodes may be in the form of an array. The elements of the array can be turned on or off on demand to create a desired spatial pattern of energy to form a desired shape of the hydrogel. For example, one or more electrodes (e.g., an array of electrodes) may be located within one or more portions of a fluid device. In another example, one or more electrodes (e.g., an array of electrodes) may be in communication (e.g., electrical communication) with one or more portions of a fluid device.

[0228] In some embodiments, the energy source is a photogenerating device. In some embodiments, the photogenerating device generates light in the range of approximately 350 nanometers to approximately 800 nanometers. In some embodiments, the photogenerating device generates light in the ranges of approximately 350 nanometers to approximately 400 nanometers, approximately 350 nanometers to approximately 450 nanometers, approximately 350 nanometers to approximately 600 nanometers, approximately 350 nanometers to approximately 800 nanometers, approximately 400 nanometers to approximately 450 nanometers, approximately 400 nanometers to approximately 600 nanometers, approximately 400 nanometers to approximately 800 nanometers, approximately 450 nanometers to approximately 600 nanometers, or approximately 600 nanometers to approximately 800 nanometers. In some embodiments, the photogenerating device generates light in the ranges of approximately 350 nanometers, approximately 400 nanometers, approximately 450 nanometers, approximately 600 nanometers, or approximately 800 nanometers. In some embodiments, the photogenerating device generates light of at least about 350 nanometers, about 400 nanometers, about 450 nanometers, or about 600 nanometers. In some embodiments, the photogenerating device generates light of up to about 400 nanometers, about 450 nanometers, about 600 nanometers, or about 800 nanometers. In some embodiments, the photogenerating device generates UV light.

[0229] In some embodiments, the mask may prevent or be configured to prevent one or more portions of the energy-emitting surface 210 of the energy source 203 from emitting energy (e.g., a non-emitting portion 204). In some embodiments, the mask may be a virtual mask (e.g., computer code or a digital system). In certain embodiments, the mask may prevent energy from being radiated to the location where a biological component is present. This may allow or permit the formation of a polymer matrix adjacent to, on, or enclosing the biological component (e.g., holding cells, proteins, DNA molecules, RNA molecules, or other target molecules in location on a fluid channel). In other embodiments, the mask may facilitate polymerization so that the polymer matrix is ​​on the biological component. In various embodiments, the mask may be a physical mask (e.g., an opaque material, a thermal shield, or an electromagnetic shield). In some embodiments, the mask (e.g., a virtual mask or a physical mask) may be generated using, or in combination with, a detector that detects or identifies the location of the biological component. In some embodiments, the detector includes a camera. In some embodiments, the detector includes a photodetector, a conductivity detector, an ultrasonic detector, an ultrasonic sensor, a piezoelectric sensor, a combination thereof, or another suitable detection device.

[0230] In some embodiments, the first surface 201 or the second surface 202 may include a detector configured to detect or detect one or more locations of one or more biological components in the fluid device (e.g., in the channel 200). In certain embodiments, the energy source 203 may include, be coupled to, or communicate with a detector configured to detect or detect the locations of biological components in the fluid device. In some embodiments, the detector may be a microscope objective lens for imaging the fluid device. In various embodiments, the mask may be generated using an image obtained from at least a portion of the fluid device. The mask may allow or permit the energy source 203 to radiate energy at or in the direction of one or more locations or positions where one or more biological components are present on or adjacent to the first surface 201. The mask may inhibit or prevent the energy source 203 from radiating energy at or in the direction of one or more locations or positions where one or more biological components are present on or adjacent to the first surface 201. In some embodiments, images may be obtained from a camera (e.g., a digital camera, a fluorescence imaging camera, etc.). In some embodiments, imaging may be bright-field imaging, phase-contrast imaging, or fluorescence imaging, or any combination thereof. In some embodiments, the camera may be coupled to, connected to, or in communication with an energy source 203. For example, a camera (not shown) may be electrically in communication with the energy source 203. In some embodiments, the energy source 203 may include a camera. In various embodiments, the energy source 203 may include a microscope (e.g., a fluorescence microscope, a confocal microscope, a lens-free imaging system, a transmission electron microscope (TEM), a scanning electron microscope (SEM), etc.). The microscope may be used to detect one or more locations of one or more biological components (e.g., in combination with a detector).

[0231] In some embodiments, an algorithm is used to determine the location of biological components or analytes based on imaging. In some embodiments, the algorithm is a supervised learning algorithm, a self-supervised learning algorithm, or an unsupervised learning algorithm. In some embodiments, the objective lens is coupled to an energy source that radiates energy to a given portion in a fluid channel.

[0232] Figure 9A shows an example of a mask including an energy masking region 910 and an energy permeable region 915. Energy from an energy source may be blocked by the energy masking region 910 to prevent the energy from forming an arbitrary polymer matrix in a portion of the fluid device (e.g., portion 920). The energy permeable region 915 may allow the energy to communicate with the fluid device to form a polymer matrix 925. Figure 9B shows another example of a mask, where the energy permeable region 935 is in the shape of a hollow cylinder (e.g., a donut). Energy masked by the masking region 930 may be prevented from energetically communicating with a portion of the fluid device (e.g., portion 940). The energy permeable region 935 may deliver energy to the fluid device to form a polymer matrix 945. The polymer matrix 945 may be in the shape of a hollow cylinder.

[0233] Figure 10 shows examples of biological components encapsulated and / or localized using a polymer matrix (i.e., indicated by white spots). In some cases, the biological component 1001 may be localized within a hollow region of the polymer matrix compartment 1002. In some other cases, the polymer matrix 1003 may be formed on the biological component 1004. In some alternative cases, the polymer matrix 1005 may localize more than one biological component. The polymer matrix 1006 of the biological compartment may encapsulate one or more biological components.

[0234] In some embodiments, the fluid device includes one or more distinct locations, one or more of which are not in fluid communication with another distinct location. In some embodiments, one or more distinct locations contain an analyte. In some embodiments, one or more distinct locations are one or more well plates. Figure 20 shows an example of a biological component 2002 encapsulated and / or localized on a well plate using a polymer matrix 2004. In some embodiments, the polymer matrix 2004 and the biological component 2002 are seeded onto a well plate 2006. In some embodiments, the biological component is introduced into the fluid device together with a polymer precursor. The polymer matrix may be formed by UV photopatterning in the absence of a physical photomask. Walls 2008 may separate individual wells. A well plate can have any number of wells. For example, a well plate can have 6, 12, 24, 48, or 96 wells. In some embodiments, each well has a diameter of about 1 mm to about 100 mm. In some embodiments, each well has a diameter of approximately 1 mm to 2 mm, approximately 1 mm to 5 mm, approximately 1 mm to 10 mm, approximately 1 mm to 20 mm, approximately 1 mm to 50 mm, approximately 1 mm to 100 mm, approximately 2 mm to 5 mm, approximately 2 mm to 10 mm, approximately 2 mm to 20 mm, approximately 2 mm to 50 mm, approximately 2 mm to 100 mm, approximately 5 mm to 10 mm, approximately 5 mm to 20 mm, approximately 5 mm to 50 mm, approximately 5 mm to 100 mm, approximately 10 mm to 20 mm, approximately 10 mm to 50 mm, approximately 10 mm to 100 mm, approximately 20 mm to 50 mm, approximately 20 mm to 100 mm, or approximately 50 mm to 100 mm.In some embodiments, each well has a diameter of approximately 1 mm, 2 mm, 5 mm, 10 mm, 20 mm, 50 mm, or 100 mm. In some embodiments, each well has a diameter of at least approximately 1 mm, 2 mm, 5 mm, 10 mm, 20 mm, or 50 mm. In some embodiments, each well has a diameter of up to approximately 2 mm, 5 mm, 10 mm, 20 mm, 50 mm, or 100 mm.

[0235] One well does not need to be in fluid communication with another well. The walls of each well 2008 can prevent fluid from moving between wells. In some cases, one or more assays may be performed on the biological components in the wells. Different assays may be performed on different wells. For example, six different assays may be performed on biological components seeded in a six-well plate.

[0236] In some embodiments, one or more distinct locations are open at the top. Figure 21 shows a gel microwell structure inside a 10 mm size well. The structure is open at the top according to some embodiments. Fluorescent beads loaded into a fluid device can also enter the well, ensuring that the well is open at the top.

[0237] Figure 3 is a flowchart illustrating the formation of a polymer matrix on or adjacent to one or more biological components according to some embodiments of the present disclosure. Process 300 may be performed manually or automatically (e.g., by a appropriately programmed computer system). In step 310, a biological sample may be placed in, introduced into, or provided to at least a portion of a fluid device. In some embodiments, the mask may then be formed or generated so that one or more portions of an energy source directed toward the biological component do not radiate (step 320). In step 330, the energy source may apply or provide energy to at least a portion of the fluid device. In some embodiments, the energy source may activate or induce a polymer precursor so that the polymer precursor forms a polymer matrix (e.g., via the energy provided by the energy source). In some embodiments, imaging of the fluid device may be performed after step 310 and before step 320 to determine or identify the locations of the biological components that will generate the mask. In some embodiments, the mask is a virtual mask. In some embodiments, the polymer matrix may form a compartment that partially or completely surrounds the biological component.

[0238] In certain cases, the energy source may be manipulated so that the polymer matrix is ​​formed in different steps. For example, the energy source may induce multiple polymer precursors so that they form open compartments (e.g., crescent-shaped or semi-cylindrical polymer matrices). The open compartments may act to trap and / or contain a portion of a biological component (e.g., a cell) or sample in a portion of a fluid device. The orientation of the energy source or fluid device may be adjusted so that additional portions of the polymer matrix are formed. These additional portions may be used in conjunction with a pre-formed semi-cylindrical polymer matrix to form one or more compartments. In other embodiments, polymer matrix compartments can be formed in at least two, three, four, five, or more matrix-forming steps.

[0239] Figures 11A and 11B illustrate examples of multi-step polymer matrix compartment generation. Figure 11A shows the first step of multi-step generation, where an open compartment (e.g., an open compartment 1101 made from a polymer matrix) may be generated to capture and / or contain a biological component (e.g., a biological component 1102). The sample containing the biological component 1102 may have a flow direction 1103 within a fluid device (e.g., a portion of a fluid device 1100). The open compartment 1101 may be formed by generating a polymer matrix using an energy source and energy adjustment unit described herein. The open compartment may intersect a portion of the flow direction 1103 of the sample in the fluid device. The polymer matrix open compartment 1101 may be oblique or perpendicular to the flow direction 1103 of the sample in the fluid device. Figure 11B shows a second step of multi-step generation, where an open compartment (e.g., open compartment 1101) is sealed or closed by forming a polymer matrix adjacent to, around, or on a biological component (e.g., biological component 1112). In some cases, in the second step, the biological component may be completely or substantially completely encapsulated by the polymer matrix (e.g., to form a closed compartment 1111). In some cases, the polymer matrix that may be formed adjacent to, around, or on the biological component localizes the biological component to a location on the fluid device 1100. Genomic material and / or proteomics material may be extracted from the localized biological component. The polymer matrix may further localize the extracted material. The fluid device may then provide a surface on which the extracted material can be sequenced. In some embodiments, the extracted material may be eluted and transferred to another device or surface for sequencing.In other embodiments, sequencing may be performed by short-read sequencing, nanopore sequencing, synthesis sequencing, in situ hybridization sequencing, any optical readout using a microscope, or any other suitable sequencing method.

[0240] One or more surfaces of a fluid device may include optical (e.g., fluorescent), mechanical, electrical, or biochemical sensing elements or sensors. Sensing elements may include fluorescent tags, enzymes, primers, oligonucleotides, or sensor molecules (e.g., biochemical sensor molecules). Sensing elements may be used to detect and / or measure pH, oxygen concentration, CO2 concentration, or any other suitable variable. Sensing elements may detect and / or measure parameters locally. For example, sensing elements may detect and / or measure pH, oxygen concentration, or CO2 concentration within a compartment surrounding a biological component (e.g., a polymer matrix shell cylinder).

[0241] System with a capture element The disclosure also provides a system comprising one or more capture elements for immobilizing and / or compartmentalizing one or more biological components. The system may include a fluid device. The fluid device may contain or include one or more biological components. Furthermore, the fluid device may contain or include one or more polymer precursors. In some embodiments, the fluid device may include a first surface (e.g., in the channels and / or chambers of the fluid device). The fluid device may include one or more capture elements. The capture elements may be capable of immobilizing or configured to immobilize at least one of one or more biological components on or adjacent to the first surface (or any preferred surface). Immobilization or coupling of a biological component to a capture element may form an immobilized biological component. The system may further include at least one energy source communicating with the fluid device. In certain embodiments, at least one energy source may be capable of supplying or providing energy to or to at least a portion of the fluid device, or may be configured to supply or to provide energy. Therefore, an energy source can activate, or cause, one or more polymer precursors (e.g., placed in a fluid device) to form at least one polymer matrix on or adjacent to an immobilized biological component. In various embodiments, the fluid device may further include a platform or stage for holding the fluid device. In some embodiments, the system may also include a sequencing device (e.g., a next-generation sequencing device) for obtaining sequencing data. The polymer matrix formed in the fluid device may be used to capture and localize the biological component. Genomic material and / or proteomics material may be extracted using the fluid device. The fluid device may then provide a surface on which the extracted material can be sequenced.In some embodiments, the extracted material may be eluted and transferred to another device or surface for sequencing. In other embodiments, sequencing may be performed by short-read sequencing, nanopore sequencing, sequencing by synthesis, sequencing by in situ hybridization, or any optical readout using a microscope.

[0242] To immobilize biological components, the fluid device may include one or more capture sites. The capture sites may include capture elements. In some embodiments, one or more capture elements or capture sites may include a pattern or be arranged in a pattern. The fluid device is 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, 30, 40, 50, 100, 200, 300, 400, 500, 600, 700, 1,000, 10,000, 10 5 , 10 6 , 10 7 , 10 8 , 10 9 , 10 10 , 10 20 The device may include a capture element, or any number between any two of the numbers referred to herein. In some embodiments, the fluid device includes 10 20 It may include more supplementary elements.

[0243] The fluid device may include channels. The fluid device may include chambers. Figure 4A shows an example of at least a portion of a channel 400 in a fluid device. One or more capture elements 411 may be disposed or positioned on a first surface 401 of the fluid device. In some cases, a second surface 402 may include one or more capture elements. The capture elements may be disposed on both surfaces or on any other suitable surface. The capture elements may contain functional groups, or may be at least partially formed by them. Some non-limiting examples of functional groups include capture reagents (e.g., pyridine carboxyaldehyde (PCA)), biotin, streptavidin, strep-tag II, linkers, or functional groups that can react with molecules (e.g., aldehydes, phosphates, silicates, esters, acids, amides, alkynes, azides, or aldehyde dithiolanes). The functional groups may be specifically coupled to the N-terminus or C-terminus of a peptide. The functional groups may be specifically coupled to amino acid side chains. The functional group may be coupled to the side chain of an amino acid (e.g., an acid of glutamic acid or aspartic acid, a thiol of cysteine, an amine of lysine, or an amide of glutamine or asparagine). The functional group may be specifically coupled to a reactive group on a particular species, e.g., a membrane-binding molecule on a cell (e.g., a glycoprotein in eukaryotic cells, or pili on the protoplasm (plasma) of prokaryotes). In some examples, the capture element may contain fibronectin. In other examples, the capture element may contain an RGD peptide. In some cases, the capture element may contain an antibody. In some examples, the functional motif may be reversibly coupled and cleaved (e.g., by using an enzyme). Figure 4B illustrates an example of a biological component 51 in contact with or coupled to the capture element 411. In some cases, a repulsive surface coating (e.g., PEG) may be used to prevent the polymer matrix from covering or trapping the biological component.

[0244] In various examples, the capture element may include physical traps, hydrodynamic traps, geometric traps, wells, electrochemical traps (e.g., trapping charged molecules), streptavidin, antibodies, aptamers, affinity binding (e.g., peptides that can bind to cell surface proteins), one or more magnetic materials (e.g., magnetic disks, magnetic arrays, or magnetic particles), dielectrophoretic traps (e.g., electrode arrays), or combinations thereof. The trap may include a polymer matrix or a hydrogel. The polymer matrix or hydrogel trap may be constructed or dismantled on demand using an energy source and / or decomposition similar to that of the polymer matrix compartments referred to herein. For example, the capture element may include wells. The wells may have a diameter of 1 μm to 50 μm. In some embodiments, the wells may have a diameter of 1 μm to 20 μm, 20 μm to 30 μm, 30 μm to 40 μm, or 40 μm to 50 μm. The wells may have a diameter greater than 50 μm. The wells may have a diameter of less than 1 μm. In some embodiments, the wells may have a depth of 0.1 μm to 100 μm. In certain embodiments, the wells may have a depth greater than 100 μm. The wells may have a depth of less than 0.1 μm. The well depths are 0.1μm~0.5μm, 0.1μm~1μm, 0.1μm~5μm, 0.1μm~10μm, 0.1μm~20μm, 0.1μm~30μm, 0.1μm~50μm, 0.1μm~100μm, 0.5μm~1μm, 0.5μm~5μm, 0.5μm~10μm, 0.5μm~20μm, 0.5μm~30μm, 0.5μm~50μm, 0.5μm~100μm, 1μm~5μm, 1μm~10μm, 1μm~ 20μm, 1μm~30μm, 1μm~50μm, 1μm~100μm, 5μm~10μm, 5μm~20μm, 5μm~30μm, 5μm~50μm, 5μm~100μm, 10μm~20μm, 10μm~30μm, 10μm~50μm, 10μm~100μm, 20μm~30μm, 20μm~50μm, 20μm~100μm, 30μm~50μm, 30μm~100μm, or 50μm~100μm may also be used.The well depth may be approximately 0.1 μm, 0.5 μm, 1 μm, 5 μm, 10 μm, 20 μm, 30 μm, 50 μm, or 100 μm. The well depth may be at least 0.1 μm, 0.5 μm, 1 μm, 5 μm, 10 μm, 20 μm, 30 μm, or 50 μm. The well depth may be up to 0.5 μm, 1 μm, 5 μm, 10 μm, 20 μm, 30 μm, 50 μm, or 100 μm.

[0245] In some embodiments, the fluid device may include a repulsive surface coating that can be used to prevent the capture of biological components at predetermined locations. Figure 12A illustrates a portion of the surface 1201 of the fluid device, which may include capture sites 1202 and repulsive sites 1203. The surface 1201 may be functionalized using a surface coating (e.g., PEG) to generate the repulsive sites 1203. The repulsive sites 1203 can prevent biological components from binding to the surface 1201 at the locations of the repulsive sites 1203, and can deliver the biological components to the capture sites 1202. In some cases, the surface 1201 may include only repulsive sites without capture sites. The repulsive sites may first localize the biological components. The polymer matrix may be formed by directing an energy source to the repulsive sites to form compartments adjacent to biological components that may be located between the repulsive sites. Figure 12B shows an example of biological components contained in predetermined locations within a fluid device. Figure 12C shows a high-magnification example of biological components contained in predetermined locations within a fluid device.

[0246] An energy source may be used to form a polymer matrix on, around, or adjacent to at least a portion of the captured biological components. In some embodiments, a mask may be used to allow or permit the energy source to direct energy toward the location or position of the captured biological components. In certain embodiments, a mask may be used to inhibit or prevent the energy source from directing energy toward the location or position of the captured biological components. The mask may form a polymer matrix surrounding, or at least partially surrounding, one or more biological components, configured to direct energy toward a predetermined or selected location. The mask may be generated at least partially based on a pattern of capture sites (e.g., a pattern of capture sites / capture elements on the surface of a fluid device). In some embodiments, the mask may be configured to prevent energy from being directed toward locations surrounding capture sites or capture elements that are not capturing or coupling biological components. In certain particular cases, for single-cell analysis, the mask may be configured to prevent energy from being radiated adjacent to locations of capture elements that are capturing or coupling two or more biological components. In some embodiments, the mask may be configured to allow or permit the emission of energy adjacent to the locations of capture elements that capture two or more biological components, for example, to enable the analysis of intercellular interactions. In certain embodiments, the mask may be a photolithography mask or another preferred mask, as described herein. In some embodiments, the system may further include a detector, for example, a detector for detecting the location of biological components, as described herein. The mask may be generated at least partially based on the detected locations of the biological components. In addition, the mask may selectively direct or supply energy from an energy source to a fluid device, as described herein.

[0247] Figure 4C illustrates an example of a method for forming a polymer matrix adjacent to (e.g., surrounding) a biological component. The polymer matrix 408 may be formed adjacent to the capture element 411. The polymer matrix 408 may be configured to hold the biological component 51 in a fixed position within or in an analysis chamber or compartment 420. The compartment 420 may be at least partially formed by the polymer matrix 408, a first surface 401, and a second surface 402, forming a chamber or at least partially sealed space (e.g., around the biological component 51) within the fluid device. In some embodiments, the polymer matrix 408 may form a compartment 420 surrounding the biological component 51. The compartment 420 may hold the biological component 51 in a fixed position. The polymer matrix 408 and / or compartment 420 may inhibit or prevent compounds associated with the biological component 51 from leaving the compartment. In some embodiments, the compounds related to the biological components may include nucleic acids (e.g., DNA or RNA), proteins, metabolites, enzymes, antibodies, combinations thereof, or any other suitable compounds or materials. In some embodiments, the surface of the polymer matrix or hydrogel may be functionalized by coupling functional groups to the polymer matrix or hydrogel. The functionalized surface of the polymer matrix inside the compartment may be coupled to a capturing element (e.g., an antibody) to capture molecules (e.g., secreted proteins) secreted by the biological components. The capturing element or captured molecules may then be read out by a sensing molecule or by a labeling method, such as fluorescent labeling. In some embodiments, the polymer matrix may be configured to allow the passage of one or more compounds related to the biological components. In some embodiments, the polymer matrix may be configured to allow the passage of reagents.The reagent may include, for example, one or more enzymes, chemicals, oligonucleotides (e.g., one or more primers having a size of less than 50 base pairs), lysozyme, proteinase K, random hexamer, polymerase, transposase, ligase, catalytic enzyme, deoxynucleotide triphosphate, buffer, cell culture medium, divalent cation, combinations thereof, or any other suitable reagent.

[0248] The pore size in a polymer matrix can be regulated using chemical reagents or by applying heat, electricity, light, or other suitable stimuli. In other words, the polymer matrix may have tunable properties (e.g., pore size). In some cases, the polymer matrix may contain thermoresponsive or temperature-responsive polymers. Thermoresponsive polymers (e.g., poly(N-isopropylacrylamide) (NIPAAM)) may undergo phase separation from the solution upon heating or cooling (e.g., the polymer exhibits a lower critical dissolution temperature (LCST) or upper critical dissolution temperature (UCST)). The polymer matrix may contain polymers that can degrade at high temperatures, for example, to control the pore size of the hydrogel or polymer matrix. Non-limiting examples of thermoresponsive polymers that can be used to form a hydrogel / polymer matrix with tunable properties include poly(N-vinylcaprolactam), poly(N-ethyloxazoline), poly(methylvinyl ether), poly(acrylic acid-co-acrylamide), or combinations thereof. Temperature changes may close or open pores in the polymer matrix to allow reagents, nucleic acid molecules, proteins, or any biomolecules or molecules smaller than the pore size to be released from the polymer matrix compartment. In some cases, the released molecules may be the target molecules. The released molecules may be collected for analysis. The pore size may decrease after the release of molecules to localize biological components and other molecules within the polymer matrix. In some cases, the remaining localized molecules may be the target molecules. For example, the pore size may be adjusted to allow protein tags (e.g., shorter DNA oligomers) to be released from the polymer matrix compartment and the fluid chamber. The protein tags may then be collected while the mRNA remains localized within the polymer matrix compartment for further analysis.

[0249] The polymer matrix may have pore sizes ranging from approximately 5 nanometers (nm) to approximately 100 nm. The polymer matrix may have pore sizes ranging from approximately 5 nm to approximately 10 nm, approximately 5 nm to approximately 20 nm, approximately 5 nm to approximately 30 nm, approximately 5 nm to approximately 40 nm, approximately 5 nm to approximately 50 nm, approximately 5 nm to approximately 60 nm, approximately 5 nm to approximately 70 nm, approximately 5 nm to approximately 80 nm, approximately 5 nm to approximately 90 nm, approximately 5 nm to approximately 100 nm, approximately 5 nm to approximately 110 nm, approximately 10 nm to approximately 20 nm, approximately 10 nm to approximately 30 nm, approximately 10 nm to approximately 40 nm, approximately 10 nm to approximately 50 nm, approximately 10 nm to approximately 60 nm, approximately 10 nm to approximately 70 nm, and approximately 10nm to about 80nm, about 10nm to about 90nm, about 10nm to about 100nm, about 10nm to about 110nm, about 20nm to about 30nm, about 20nm to about 40nm, about 20nm to about 50nm, about 20nm to about 60nm, about 20nm to about 70nm, approximately 20nm to approximately 80nm, approximately 20nm to approximately 90nm, approximately 20nm to approximately 100nm, approximately 20nm to approximately 110nm, approximately 30nm to approximately 40nm, approximately 30nm to approximately 50nm, approximately 30nm to approximately 60nm, approximately 30nm to approximately 70nm, approximately 30nm to about 80nm, about 30nm to about 90nm, about 30nm to about 100nm, about 30nm to about 110nm, about 40nm to about 50nm, about 40nm to about 60nm, about 40nm to about 70nm, about 40nm to about 80nm, about 40nm to about 90nm, about 40nm to about 100nm, about 40nm to about 110nm, about 50nm to about 60nm, about 50nm to about 70nm, about 50nm to about 80nm, about 50nm to about 90nm, about 50nm to about 100nm, about 50nm to about 110nm The polymer matrix may have pore sizes of approximately 60nm to 70nm, 60nm to 80nm, 60nm to 90nm, 60nm to 100nm, 60nm to 110nm, 70nm to 80nm, 70nm to 90nm, 70nm to 100nm, 70nm to 110nm, 80nm to 90nm, 80nm to 100nm, 80nm to 110nm, 90nm to 100nm, 90nm to 110nm, or 100nm to 110nm. The polymer matrix may have pore sizes of approximately 5nm, 10nm, 20nm, 30nm, 40nm, 50nm, 60nm, 70nm, 80nm, 90nm, 100nm, or 110nm.The polymer matrix may have pore sizes of at least about 5 nm, about 10 nm, about 20 nm, about 30 nm, about 40 nm, about 50 nm, about 60 nm, about 70 nm, about 80 nm, about 90 nm, about 100 nm, or less. The polymer matrix may have pore sizes of up to about 10 nm, about 20 nm, about 30 nm, about 40 nm, about 50 nm, about 60 nm, about 70 nm, about 80 nm, about 90 nm, about 100 nm, about 110 nm, or larger.

[0250] The polymer matrix may be degradable. In some embodiments, the degradable polymer matrix may be depolymerized (e.g., into polymer precursors). At least a portion of the polymer precursors from the degraded polymer matrix may be reused to form another polymer matrix. The polymer matrix may be depolymerized using energy sources provided herein. In some embodiments, by depolymerizing the polymer matrix, compartments surrounding biological components may be dismantled (e.g., to release the biological components). In some embodiments, releasing biological components may also include releasing the biological components from the captured element. That is, the biological components may be detached from the captured element or decoupled. In some embodiments, the captured element or a portion of the captured element may be cleaved using a chemical compound (e.g., agarase, dextranase, metalloproteinase, or other enzyme) or by providing energy to the captured site or element using an energy source (e.g., photomediated decomposition). In some cases, the captured element may be cleaved using hydrolysis, ester hydrolysis, enzymatic hydrolysis, reversible click reaction, or photodegradation cleavage. In some cases, the capture element may be decoupled from the surface or biological component by applying a physical force (e.g., sonication, agitation of a fluid device, etc.). In some cases, the capture element may contain agarose that can be degraded or cleaved using agarase. In some cases, the capture element may contain dextran that can be degraded or cleaved using dextranase. In some cases, the capture element may contain a metalloproteinase (MMP)-degradable peptide. In certain specific cases, the biological component may be decoupled from the capture element by chemical means (e.g., using digestive enzymes to cleave the bond) or by physical means (e.g., using an energy source to provide heat, microwaves, electromagnetic waves, electromagnetic fields, sound waves, etc.).

[0251] In some embodiments, the polymer matrix may be decomposed or depolymerized by contacting it with a cleavage mix. In some embodiments, the cleavage mix may contain dithiothreitol (DTT), β-mercaptoethanol, glutathione, tris(2-carboxyethyl)phosphine (TCEP), tris(3-hydroxypropyl)phosphine (THP), or a combination thereof. In some embodiments, heat may be directed to the polymer matrix to decompose or depolymerize it. In some embodiments, the polymer matrix may be heated to at least 90°C. In some embodiments, the polymer matrix may be heated to 80°C-100°C, 90°C-110°C, 110°C-120°C, 120°C-180°C, or 180°C-250°C. The polymer matrix may be decomposed or depolymerized by directing a wavelength of light suitable for cleaving a photocleavable crosslinking agent that crosslinks the polymer matrix to the polymer matrix. In some embodiments, the fluid device may include a photoactive compound (e.g., a photoacid generator or a photobase generator) that can decompose or depolymerize the polymer matrix upon exposure to light energy (e.g., a wavelength of light).

[0252] The polymer matrix may contain a hydrogel. In some embodiments, the polymer matrix may form a wall, which may be coupled to a first surface and / or a second surface. In some embodiments, the wall may be a hydrogel wall. The polymer matrix wall or hydrogel wall may have a thickness of 1 μm to 250 μm. Polymer matrix walls or hydrogel walls are available in the following thicknesses: 1μm-5μm, 1μm-10μm, 1μm-20μm, 1μm-30μm, 1μm-40μm, 1μm-50μm, 1μm-100μm, 1μm-150μm, 1μm-250μm, 5μm-10μm, 5μm-20μm, 5μm-30μm, 5μm-40μm, 5μm-50μm, 5μm-100μm, 5μm-150μm, 5μm-250μm, 10μm-20μm, 10μm-30μm, 10μm-40μm, 10μm-50μm, 10μm-100μm, 10μm-150μm, and 10μm-2 It may have a thickness of 50 μm, 20 μm to 30 μm, 20 μm to 40 μm, 20 μm to 50 μm, 20 μm to 100 μm, 20 μm to 150 μm, 20 μm to 250 μm, 30 μm to 40 μm, 30 μm to 50 μm, 30 μm to 100 μm, 30 μm to 150 μm, 30 μm to 250 μm, 40 μm to 50 μm, 40 μm to 100 μm, 40 μm to 150 μm, 40 μm to 250 μm, 50 μm to 100 μm, 50 μm to 150 μm, 50 μm to 250 μm, 100 μm to 150 μm, 100 μm to 250 μm, or 150 μm to 250 μm. The polymer matrix wall or hydrogel wall may have a thickness of approximately 1 μm, approximately 5 μm, approximately 10 μm, approximately 20 μm, approximately 30 μm, approximately 40 μm, approximately 50 μm, approximately 100 μm, approximately 150 μm, or approximately 250 μm. The polymer matrix wall or hydrogel wall may have a thickness of at least 1 μm, 5 μm, 10 μm, 20 μm, 30 μm, 40 μm, 50 μm, 100 μm, 150 μm, 250 μm, or greater. The polymer matrix wall or hydrogel wall may have a maximum thickness of 5 μm, 10 μm, 20 μm, 30 μm, 40 μm, 50 μm, 100 μm, 150 μm, or 250 μm.The polymer matrix wall or hydrogel wall may have a thickness of less than 1 μm.

[0253] In some embodiments, the hydrogel is made from polyethylene glycol (PEG)-thiol, PEG-acrylate, acrylamide, N,N'-bis(acryloyl)cystamine, PEG, polypropylene oxide (PPO), polyacrylic acid, poly(hydroxyethyl methacrylate) (PHEMA), poly(methyl methacrylate) (PMMA), poly(N-isopropylacrylamide) (PNIPAAm), poly(lactic acid) (PLA), poly(lactic acid-co-glycolic acid) (PLGA), polycaprolactone (PCL), poly(vinyl sulfonic acid) (PVSA), poly(L-aspartic acid), poly(L-glutamic acid), polylysine, and agar. The hydrogel may contain agarose, alginate, heparin, alginate sulfate, dextran sulfate, hyaluronan, pectin, carrageenan, gelatin, chitosan, cellulose, collagen, bisacrylamide, diacrylate, diallylamine, triallylamine, divinyl sulfone, diethylene glycol diallyl ether, ethylene glycol diacrylate, polymethylene glycol diacrylate, polyethylene glycol diacrylate, trimethylopropane trimethacrylate, ethoxylated trimethylol triacrylate, ethoxylated pentaerythritol tetraacrylate, or combinations or mixtures thereof. In some embodiments, the hydrogel may contain PEG-thiol / PEG-acrylate, acrylamide / N,N'-bis(acryloyl)cystamine (BACy), or PEG / PPO. For example, the PEG molecule may contain a multi-armed PEG derivative having a thiol group at each end of an arm connected to a single pentaerythritol core. Reactive free thiols, SHs, sulfhydryls, or mercapto groups may selectively react with microfluidic surfaces containing gold, silver, etc. (e.g., maleimide or transition metal surfaces). PEG-SH can be oxidized in air to form SS disulfide (disulfide) bonds, which can be reversed with a reducing agent to form reversible PEGylation or PEG hydrogels.

[0254] As discussed, the polymer matrix (e.g., hydrogel) may be sufficiently porous to allow the passage of reagents (e.g., enzymes, reagents, small molecules, antibodies) while preventing the passage of captured biological components (e.g., DNA, RNA, proteins, cells, etc.) or compounds associated with biological components (e.g., DNA, RNA, antibodies, secreted compounds from cells, etc.). In some embodiments, the reagent may include an enzyme or a primer having a size of less than 50 base pairs (bp). The primer may have a size of 5pb to 50bp. In some embodiments, the primer may have a size of 5pb to 10bp, 10bp to 20bp, 20bp to 30bp, 30bp to 40bp, or 40bp to 50bp. In certain embodiments, the primer may have a size greater than 50bp. In various embodiments, the primer may have a size of less than 5bp.

[0255] In certain embodiments, the first, second, or both surfaces of a channel in a fluid device may be functionalized as described herein. The surfaces of the fluid device (e.g., the first, second, and third surfaces) may contain compounds configured to bind to biological components (e.g., captured biological components). In some embodiments, the surfaces of the fluid device (e.g., the first, second, and third surfaces) may contain one or more barcodes. One or more surfaces may contain oligos that form DNA clusters for sequencing. In some cases, one or more surfaces may contain one or more nanopore readers for direct DNA and / or RNA readout. One or more surfaces may contain nanowells for capturing single RNA molecules and / or single DNA molecules, or for containing DNA / RNA libraries. In some alternative cases, one or more surfaces may contain patterned hydrophobic / hydrophilic properties for selective arrangement of DNA nanoballs. Nanoballs can be generated by circulating and amplifying DNA libraries from DNA / RNA molecules.

[0256] One or more surfaces of a fluid device may include optical (e.g., fluorescent), mechanical, electrical, or biochemical sensing elements or sensors. Sensing elements may include fluorescent tags, enzymes, primers, oligonucleotides, or sensor molecules (e.g., biochemical sensor molecules). Sensing elements may be used to detect and / or measure pH, oxygen concentration, CO2 concentration, or any other suitable variable. Sensing elements may detect and / or measure parameters locally. For example, sensing elements may detect and / or measure pH, oxygen concentration, or CO2 concentration within a compartment (e.g., a polymer matrix shell cylinder) surrounding or encapsulating a biological component.

[0257] Multilayer system A system for analyzing biological components is also provided herein, comprising at least one flow channel (e.g., a first layer or upper layer) and an analysis channel (e.g., a second layer or lower layer). The system may include a fluid device comprising the flow channel, the analysis channel, and a layer or wall disposed between the flow channel and the analysis channel. The system may include at least one energy source communicating with the fluid device, as described herein. The analysis channel may be disposed adjacent to the flow channel, wherein at least one flow-inhibiting element may be disposed within the flow channel to inhibit or stop the flow of biological components in the flow channel. The layer disposed between the flow channel and the analysis channel may include at least one sealable opening disposed in or adjacent to the at least one flow-inhibiting element. One or more biological components may be stopped or trapped adjacent to the sealable opening. The at least one sealable opening may be configured to allow the passage of one or more biological components. For example, the sealable opening may be configured to allow the passage of one or more biological components from the flow channel to the analysis channel. At least one energy source may be in communication with the analytical channel. Furthermore, at least one energy source may form a polymer matrix within the analytical channel, or may be configured to form one.

[0258] As described herein, in some embodiments, the fluid device may include a microfluidic device or a nanofluidic device. In certain embodiments, the fluid device may be used for nucleic acid sequencing. In some cases, the fluid device may include a nucleic acid sequencing flow cell. In other cases, the sequencing may include short-read sequencing, nanopore sequencing, sequencing by synthesis, sequencing by in situ hybridization, sequencing by collection of any optical readout, or any other suitable sequencing method. As described herein, the biological components may include cells, cell lysates, nucleic acids, microbiomes, proteins, cell mixtures, spatially linked biological components, metabolites, combinations thereof, or any other suitable biological components. In some cases, the cell mixture may include two or more different cell types. For example, the cell mixture may include a first cell type and a second cell type. In some cases, the cell mixture may include two, three, four, five, six, seven, eight, nine, ten, or more cell types. The cells may be mammalian cells (e.g., human cells), fungal cells, bacterial cells, tumor spheroids, combinations thereof, or any other suitable cells. In some cases, the biological components may include tumor spheroids or spatially linked biological components (or samples).

[0259] In some cases, nucleic acids may contain at least 100 bases or base pairs. In certain embodiments, nucleic acids include DNA or RNA. DNA may be at least 100 bp in length. In some embodiments, DNA may contain at least 50 bp, 100 bp, 200 bp, 300 bp, 400 bp, 500 bp, 600 bp, 700 bp, 800 bp, 900 bp, 1,000 bp, 10 kilobase pairs (kbp), 100 kbp, 1 megabase pair (Mbp), 100 Mbp, 1 gigabase pair (Gbp), 10 Gbp, 100 Gbp, or more base pairs. Biological components may include DNA molecules containing any number of base pairs between the numbers mentioned herein. For example, DNA may contain 50 bp to 1,000 bp, 300 bp to 10 kbp, or 1,000 bp to 10 Gbp. RNA may be dsRNA. dsRNA may contain at least 50 bp, 100 bp, 200 bp, 300 bp, 400 bp, 500 bp, 600 bp, 700 bp, 800 bp, 900 bp, 1,000 bp, 10 kbp, or 100 kbp. Biological components may contain dsRNA molecules containing any number of base pairs between the numbers mentioned herein. For example, dsRNA may contain 50 bp to 1,000 bp, 300 bp to 10 kbp, or 1,000 bp to 100 kbp. RNA may be ssRNA. ssRNA may contain at least 50 nucleotides to 100,000 nt. ssRNA may contain 50nt-100nt, 50nt-1,000nt, 50nt-10,000nt, 50nt-100,000nt, 100nt-1,000nt, 100nt-10,000nt, 100nt-100,000nt, 1,000nt-10,000nt, 1,000nt-100,000nt, or 10,000nt-100,000nt. In some cases, ssRNA may be less than 50 nucleotides in length. ssRNA may also be longer than 100,000 nucleotides.

[0260] In some embodiments, the flow channel or a portion thereof may be parallel or substantially parallel to the analysis channel or a portion thereof. In some embodiments, the flow channel may be removablely coupled to the analysis channel. For example, the user may remove the flow channel from the analysis channel. Thus, a portion of the fluid device containing the analysis channel may be used to perform various analyses or experiments. The portion of the fluid device containing the analysis channel, along with the portion of the fluid device containing the flow channel that is removed, may be easily accessible by, for example, a detector, camera, or other device for analyzing biological components within the analysis channel.

[0261] In some cases, the analytical channel may include a polymer matrix structure for capturing or trapping biological components or molecules or compounds produced by biological components (e.g., before introducing the biological components into the fluid device). For example, the user may obtain an analytical channel that includes a polymer matrix structure; that is, the user does not have to form the polymer matrix structure. In various cases, the analytical channel may be configured to include a polymer matrix structure for capturing or trapping biological components or molecules or compounds produced by biological components. For example, in such embodiments, one or more polymer matrix structures may be formed in the analytical channel after introducing the biological components into the fluid device and the analytical channel. The analytical channel may be configured for a screening process, library preparation, or another preferred process. In some embodiments, the screening process may be for drug screening, antibiotic screening, culture condition screening, or CRISPR screening. In certain cases, multiple samples may be placed in multiple channels. Multiple samples may be screened for various conditions in other signal-containing channels.

[0262] The sealable opening may be configured to transition from a sealed state to an open state. For example, the sealable opening may contain a heat-sensitive polymer that, for example, can melt when heated, thereby opening the sealable opening. In some cases, the passage of biological components through the sealable opening may be inhibited in the sealed state. In certain cases, the passage of biological components through the sealable opening may be permitted in the open state. In some cases, the sealable opening may be sealed with an agarose gel, a temperature-soluble polymer, an N-isopropylacrylamide (NIPAAm) polymer, a wax compound, an alginate, or any other suitable compound or material.

[0263] Figures 6A and 6B show a portion of a fluid device configured to trap biological components 50. The fluid device may include a flow channel or chamber 651, an analysis channel or chamber 652, and a layer or wall 653 positioned between at least a portion of the flow channel 651 and the analysis channel 652. The layer 653 may include one or more sealable openings or openings 654. In addition, one or more flow habitat elements 655 may be able to obstruct or prevent the biological components 50 from flowing along the flow channel 611, or may be configured to obstruct or prevent it. Flow obstruction elements 655 may be configured adjacent to a sealable opening 654 to stop or trap the biological components 50. As described herein, a sealable opening 614 may be configured to transition from a sealed state (e.g., closed state) to an open state or configuration. Figure 6A shows an example of a sealable opening 614 in a sealed state. Figure 6B shows an example of a sealable opening 654 in an open state. During the transition from a sealed to an open state, the sealable opening 654 may allow or permit the passage of biological components 50 from at least a portion of the flow channel 651 to at least a portion of the analysis channel 652. In certain examples, the analysis channel 652 may be installed or configured to be installed below the flow channel 651 so as to allow the biological components 50 to move from the flow channel 651 to the analysis channel 652 by a provided force (e.g., gravity, a high-pressure pulse by pressurizing the flow in the flow channel, and the generation of negative pressure in the analysis channel). In some embodiments, a fluid device may be spun or centrifuged to position one or more biological components from the flow channel to the analysis channel. Reagents may be placed in or pass through at least a portion of the analysis channel 652 for, for example, to perform an analysis or experiment provided herein.

[0264] As shown in Figure 6A, the flow obstruction element 655 may be positioned within at least a portion of the flow channel 651 to obstruct or prevent the flow of biological components (e.g., biological components 50) in the flow channel 651. The flow obstruction element 655 may be configured to capture or trap the biological components 50 in at least a portion of the flow channel 651. In some cases, the flow obstruction element 655 may extend from the surface of the flow channel 651 (e.g., surface 669). In some cases, surface 669 may be positioned opposite a flow channel surface 661 adjacent to layer 653.

[0265] In various cases, the analysis channel 652 may include a surface 659 positioned opposite the analysis channel surface 663, which is adjacent to or is the surface of layer 653. The analysis channel 652 may include one or more polymer matrices 656. The analysis channel 652 may include one or more polymer precursors. For example, one or more polymer precursors may be located in at least a portion of the analysis channel 652. One or more polymer matrices 656 may be formed in the analysis channel 602 using an energy source that provides energy to one or more polymer precursors. The energy source may be in optical, electrochemical, electromagnetic, thermal, or microwave communication with a fluid device or the analysis channel 652. In some cases, the energy source may be a photogenerating device, a thermogenerating device, an electrochemical generating device, an electrode, a microwave device, or a combination thereof. The energy source may selectively provide energy to the analysis channel 652 to form the polymer matrix at a predetermined location. A spatial energy modulating element may be used to selectively provide energy to the analysis channel 652.

[0266] In some cases, the spatial energy modulating element may include a photolithography mask, a DMD system, or other suitable mask. One or more polymer matrices 656 may be formed before the transition of the sealable opening 654 to an open state (e.g., as shown in Figure 6A). For example, the polymer matrix may be formed with and aligned with the sealable opening so that the biological components 650 held by the inhibitory element 655 can be directed to the compartment 620 when the sealable opening 654 is opened (e.g., fall by gravity or fluid pressure). One or more polymer matrices 656 may be formed after the transition of the sealable opening 654 to an open state (e.g., as shown in Figure 6B). One or more polymer matrices 656 may form an analysis chamber or compartment 620 as described herein.

[0267] Figures 7A and 7B show top views of the fluid device. The flow-blocking channel 675 may be configured to block the flow of the biological component 20 along the flow channel 651. The flow of the fluid (e.g., the fluid containing the biological component) through the flow channel 651 and the flow-blocking channel 651 may trap or stop the biological component 20 at the opening of the flow-blocking channel 675, as shown in Figure 7A. As shown, the dimensions (e.g., width) of the flow-blocking channel 675 may be too small or too narrow to allow or permit the passage of the biological component 20 through the flow-blocking channel 675. As shown in Figure 7B, the polymer matrix 676 may be formed on or adjacent to (e.g., surrounding) the biological component 20. In some cases, the polymer matrix may surround at least a portion of the biological component. The fluid device in Figures 7A and 7B may be a single-layer fluid device; that is, the polymer matrix may be formed in the flow channel 651. As shown in the illustration, the path of the flow channel 651 may be winding. For example, the flow channel 651 may include one or more curves. In some embodiments, the path of the flow channel may be straight, substantially straight, a zigzag pattern, or any other preferred shape.

[0268] In certain embodiments, the fluid device in Figures 7A and 7B may include two or more layers. For example, the fluid device may include a flow channel and an analysis channel (similar to the system shown in Figures 6A and 6B). Furthermore, a sealable opening may be located in part of or adjacent to the flow-blocking channel. In such embodiments, biological components may be moved through the sealable opening to the analysis channel (e.g., located adjacent to or below the flow channel) as described herein. In some cases, the analysis channel may accept two or more biological components. For example, the analysis channel may accept two, three, four, five, six, seven, eight, nine, ten, or more biological components.

[0269] Figure 8 shows an example of a fluid device containing, or configured for, multiple reagents and / or analytes (R1, R2, R3, and R4). The fluid device may include a first flow channel 851a for receiving one or more biological components from a first sample. The first flow channel 851a may allow or permit the flow or passage of one or more biological components from the first sample. Furthermore, the first flow channel 851a may allow or permit the flow or passage of one or more polymer precursors. The fluid device may include a second flow channel 851b for receiving one or more biological components from a second sample. The second flow channel 851b may allow or permit the flow or passage of one or more biological components from a second sample. Furthermore, the second flow channel 851b may allow or permit the flow or passage of one or more biological components from the second sample.

[0270] The first flow channel 851a and / or the second flow channel 851b may include a plurality of inhibitory elements (e.g., inhibitory element 855). A biological component (e.g., biological component 50) may be trapped or localized by the inhibitory elements 855. As described herein, the first flow channel 851a and / or the second flow channel 851b may include one or more sealable openings located in or adjacent to one or more inhibitory elements 855 that can be opened (e.g., transition from a sealed state to an open state) to allow the biological component to move into the first analysis channel 852a or the second analysis channel 852b. The first and second flow channels 851a, 851b may be located above the first and second analysis channels 852a, 852b (e.g., in upper and lower layers similar to the fluid device illustrated in Figures 6A and 6B). A polymer matrix 856 may be formed surrounding the biological component 50. The polymer matrix 856 may partially surround the biological component 50. The polymer matrix 856 may form a compartment or analytical chamber 820 that localizes the biological component 50 within at least a portion of the analytical channel (e.g., analytical channels 851a, 851b).

[0271] The first analytical channel 852a may contain one or more reagents and / or analytes different from the one or more reagents and / or analytes in the second analytical channel 852b. The first analytical channel 852a may contain the same one or more reagents and / or analytes as the one or more reagents and / or analytes in the second analytical channel 852b. In some cases, the fluid device may contain at least 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 25, 50, or more flow channels. In certain particular cases, the fluid device may contain at least 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 25, 50, or more analytical channels. The fluid device may analyze multiple biological components in parallel. Multiple biological components may be exposed to one or more different reagents and / or analytes as provided herein. Such a configuration (for example, as shown in Figure 8) may allow for the analysis of multiple biological components in one or more samples under various conditions provided by different reagents and / or analytes. The fluid device shown in Figure 8 may be used in a screening process. The screening process may be for drug screening, antibiotic screening, culture condition screening, or CRISPR screening. The screening process may be carried out in a combined manner. For example, multiple samples may be loaded (e.g., in parallel) into multiple flow channels that can be screened against multiple conditions in multiple analytical channels.

[0272] The first and / or second samples may be homogeneous or heterogeneous. For example, one or more biological components in the first sample may be the same or different. The first sample may be different from the second sample. In some cases, the biological components may be released from the compartment or analysis chamber 820 by selectively degrading the polymer matrix, as described herein. In other words, the polymer matrix may be degraded "on demand" (e.g., as directed by the user or computer). In various embodiments, degradation may be achieved by the use of localized stimuli. In certain embodiments, degradation may be achieved by the use of heat, light, electrochemical reactions, or a combination of some of these. The released biological components may be collected using an exit channel (e.g., exit channel 881a or 881b).

[0273] As described in relation to the fluid device in Figures 6A and 6B, the layer may be positioned between the flow channels 851a, 851b and the analysis channels 852a, 852b. The analysis channel surface adjacent to the layer (e.g., similar to surface 661 shown in Figure 6A), the analysis channel surface facing the layer (e.g., similar to surface 659 shown in Figure 6A), or both, may contain one or more barcodes as described herein.

[0274] In some cases, channels (e.g., channels 100, 200, 400) and / or analytical channels (e.g., analytical channels 652, 852a, 852b) may contain molecules in addition to, or instead of, one or more barcodes. For example, the surface of one or more channels and / or analytical channels may contain optical (e.g., fluorescent), mechanical, electrical, or biochemical sensing elements or sensors. Sensing elements may include fluorescent tags, enzymes, primers, oligonucleotides, or sensor molecules (e.g., biochemical sensor molecules). Sensing elements may be used to detect and / or measure pH, oxygen concentration, CO2 concentration, or any other suitable variable. Sensing elements may detect and / or measure parameters locally. For example, sensing elements may detect and / or measure pH, oxygen concentration, or CO2 concentration within a compartment surrounding a biological component (e.g., a polymer matrix shell cylinder).

[0275] Figure 16A shows a portion of any of the systems provided herein, including an energy source 1610 (e.g., a light source) and a spatial energy modulating element (e.g., a mask) 1630. In some embodiments, the energy source 1610 may be coupled to an objective lens (e.g., a microscope objective lens or lens). The energy source 1610 may radiate energy as electromagnetic waves (e.g., microwaves, light, heat, etc.). In some embodiments, the radiated energy 1620 may or may be sufficient to form a polymer matrix 1660 within at least a portion of the channel 1665. In some embodiments, the energy source 1610 is configured to radiate energy only towards a specific targeted portion of the channel 1665. In some embodiments, this can be achieved by using a spatial energy modulating element or mask 1630. For example, a non-radiating portion 1650 of the spatial energy modulating element 1630 may inhibit or prevent the direction of energy to one or more locations on the channel 1665 (e.g., locations of biological components). The radiating portion 1640 may allow energy 1620 radiated by the energy source 1610 to come into contact with a targeted location on the channel 1665. In some embodiments, the mask 1630, including the radiating portion 1640 and the non-radiating portion 1650, may include physical components (e.g., opaque materials, thermal shields, or electromagnetic shields). In other embodiments, the mask 1630, the radiating portion 1640, the non-radiating portion 1650, or a combination of some of them, may include digital components (e.g., computer code or a digital system that operates the electrodes). For example, a digital mask or virtual mask may generate spatially regulated energy (e.g., light, electric current, etc.) on one or more electrodes or arrays of electrodes to form a patterned polymer matrix 1660.

[0276] Continuing with Figure 16A, in some cases, the polymer matrix 1660, or at least a portion of the polymer matrix 1660, may surround or substantially surround the biological component 1670. In some embodiments, the polymer matrix 1660 may encapsulate the biological component 1670 by itself or in conjunction with channels and other surfaces.

[0277] In some embodiments, either the channel or the surface of the analytical channel may be configured to have functional groups as described herein. In certain embodiments, the system may further include a detector for detecting (or identifying) biological components, one or more barcodes, or combinations thereof, as described herein. In various embodiments, the system may further include a platform or stage configured to hold a fluid device. In some embodiments, the system may further include a sequencing device. In some cases, the system may further include a space energy modulator for selectively supplying energy to the fluid device, as described herein.

[0278] Figure 16B shows a top view of the channel provided herein. Energy radiated from an energy source can be spatially regulated as described herein to form a polymer matrix 1660. In some examples, the polymer matrix 1660 can encapsulate biological components 1670. In certain examples, the biological components can be encapsulated in and / or together with the polymer matrix 1660 by the channel walls.

[0279] Figures 17A–17E illustrate examples of workflows in which energy is applied to one of the fluid devices described herein to form a polymer matrix. Figure 17A shows a portion of the channel 1730 of the fluid device, a first biological component 1720, a second biological component 1725, an energy source (e.g., a light source) 1700, and an energy modulating element (e.g., a mask or localized mask) 1710. In some embodiments, the energy source 1700 may be positioned adjacent to or near the channel 1730 so that sufficient energy to induce hydrogel polymerization can be directed from the energy source 1700 to at least a portion of the channel 1730. In some embodiments, a spatial energy modulating element 1710 may be used to selectively allow energy from the energy source 1700 to communicate with at least a portion of the fluid device. Biological samples or biological components 1720, 1725 may be present within the channel 1730. Figure 17B shows the first step of hydrogel formation at a first location (e.g., adjacent to the first biological component 1720). The energy source 1700 can emit enough energy 1750 to form the polymer matrix 1760 in the channel 1730 adjacent to the first biological component 1720.

[0280] In some embodiments, energy can be guided by or pass through the spatial energy modulator 1710 as described herein. In some examples, a polymer matrix can be formed at locations in the channel corresponding to the radiating portion of the spatial energy modulator 1710. Figure 17C shows a second step of hydrogel formation in channel 1730, where the relative positions of the energy source 1700 and channel 1730 can change. In some embodiments, the fluid device may be mounted on a movable stage. In certain embodiments, the energy source 1700 can be movable or coupled to a movable stage. In various embodiments, the mask 1710 can be movable or coupled to a movable stage. In some embodiments, some combinations of the energy source 1700, mask 1710, and fluid device can be configured to be movable relative to each other. In some embodiments, the spatial energy modulator 1710 can be a virtual spatial energy modulator, and the position of the radiating area can change digitally.

[0281] Figure 17D shows a third step, where energy passing from the energy source 1700 through the energy modulating element (or mask) 1710 can form a polymer matrix at a second location (e.g., adjacent to the second biological component 1765). In some embodiments, the energy source 1700, the mask 1710, and the fluid device can move relative to each other, as described herein, and use the energy from the energy source to form different hydrogel patterns within the channel 1730, including multiple hydrogel matrices (e.g., hydrogel matrices 1765, 1766, 1767, and 1768) (Figure 17E).

[0282] Methods for analyzing biological components Methods for analyzing biological components are also provided herein. The method may include the steps of introducing a biological component into a fluid device, and forming a polymer matrix on or adjacent to the biological component. The method may further include the step of coupling the biological component to one or more capture elements disposed on the surface of the fluid device (e.g., a first surface) to obtain a coupled biological component. Thus, the polymer matrix can be formed on or adjacent to the coupled biological component.

[0283] Figure 5 is a flowchart of an example of a method for analyzing biological components using a system including a fluid device disclosed herein. Method 500 may include a step 510 of providing and / or introducing a biological sample to the fluid device, which may or may be suspected of containing biological components. In a capture step 520, a capture element in the fluid device may capture and / or couple the biological components, for example, so that the biological components are immobilized. In some cases, the immobilization of the biological components may be performed by using a repulsive surface coating to contain the biological components within a portion of a surface that does not have a repulsive surface, as described herein. In some cases, the capture step may be omitted, and the biological components may be randomly distributed across the surface before step 530. In step 530, a spatial energy modulating element (e.g., a mask) may be selectively applied to provide energy from an energy source to the fluid device to form one or more polymer matrices. In some embodiments, the mask may be configured to selectively supply energy adjacent to a biological component to form a polymer matrix adjacent to the biological component. In certain embodiments, the mask may be configured to selectively supply energy onto a biological component to form a polymer matrix that encapsulates at least a portion of the biological component. The mask may be applied based at least partially on the location of the biological component. Method 500 may further include the step of forming or generating a predefined or predetermined pattern of capture sites / capture elements on one or more surfaces of a fluid device (e.g., a first surface, a second surface, a third surface, etc.) prior to step 510. In some cases, the predefined or predetermined pattern of capture sites / capture elements may be generated on the fluid device prior to Method 500. In some embodiments, each capture site may be configured to have one or more capturing elements as described herein.

[0284] In some embodiments, a detector may be used to detect the location or position of the biological components, as provided herein. A mask may then be generated based at least partially on the detected location or position of the biological components. In step 540, the generated mask may be used in combination with an energy source to selectively apply energy to the fluid device and / or the biological sample introduced into the fluid device in step 510. In certain embodiments, the mask may be a photolithography mask or another suitable mask. In step 550, a polymer matrix may be formed by applying sufficient energy (e.g., from an energy source) to polymerize the polymer precursor in the fluid device and / or the biological sample introduced into the fluid device in step 510. The energy may include electrochemical energy, electromagnetic energy, thermal energy, microwave energy, or any other suitable energy. In some embodiments, the energy may be light energy, as discussed herein.

[0285] In certain embodiments, the polymer matrix may be formed adjacent to the biological component. In some embodiments, the polymer matrix may be formed on the biological component (e.g., to encapsulate the biological component). The polymer matrix may be formed between two biological components to prevent contact (e.g., physical contact) between the two biological components. In various embodiments, at least a portion of the biological component may be surrounded by the polymer matrix. In some embodiments, one or more biological components in a biological sample may be surrounded by a polymer matrix so that two biological components can be separated from each other by the polymer matrix (e.g., one or more polymer matrix walls). The biological component may be encapsulated by the polymer matrix. In certain embodiments, the polymer matrix may include a hydrogel. In various embodiments, the polymer matrix may form a compartment around the biological component (e.g., by surrounding the biological component) to form an analysis chamber.

[0286] As described elsewhere in this specification, one or more assays may be performed on biological components in an analytical chamber. One or more biological components in a biological sample may be captured, and the analytical chamber may be formed adjacent to and / or surrounding each of the captured biological components. The order of steps or actions of the methods described in relation to the disclosed embodiments may be changed. Therefore, any order in the drawings or detailed description is for illustrative purposes only and does not imply a required order unless otherwise specified.

[0287] In some embodiments, one or more functional assays may be performed on or against biological components in a polymer matrix. In some embodiments, functional assays are used to evaluate cell viability, cell morphology, cell secretion, cell response, cell-cell interactions, or any combination thereof. In some embodiments, colorimetric assays or fluorescence assays may be performed or against. In some embodiments, functional assays are performed using bright-field phase imaging or fluorescence imaging of the analyte. In some embodiments, cells may be lysed, and functional assays will be performed against their components. One or more individual components may be localized (e.g., encapsulated) within the fluid device, and the localized components may be exposed to one or more reagents and / or washing solutions during and / or between analyses, so that multiple assays can be performed within a compartment (e.g., simultaneously, substantially simultaneously, sequentially, etc.). Different assays may be performed at different locations in the fluid device, for example, to test the effects of different processing conditions.

[0288] In some embodiments, one or more omics assays may be performed to characterize and quantify biological components or their components. The omics assays may be proteomics assays, transcriptomics assays, genomics assays, or epigenomics assays, or any combination thereof. In some embodiments, one or more omics assays are multi-omics assays.

[0289] Methods for obtaining a transcriptome of a biological component are also provided herein. The method may include the step of coupling the biological component to a capture element placed in a fluid device to obtain a coupled biological component. The method may include the step of forming an analysis chamber by forming a polymer matrix on or adjacent to the coupled biological component. The method may further include the step of carrying out or performing one or more reactions in the analysis chamber to obtain a transcriptome of the biological component. The biological component may remain in the analysis chamber during the performance of one or more reactions. In some embodiments, the biological component does not need to be coupled to a capture element placed in a fluid device. For example, the biological component can be localized by forming a polymer matrix on or adjacent to the biological component without coupling a biological device to a capture element. The transcriptome may be captured by a surface-mounted target capture probe. The capture probe may include a barcode that decodes the location of the biological component and distinguishes the biological component from the source. Furthermore, biochemical treatments can be performed on the captured transcriptome of biological components, using sequencing or hybridization to convert them into DNA and determine their sequence.

[0290] The method may further include the step of directing energy from an energy source to at least a portion of a fluid device to form a polymer matrix on or adjacent to a biological component to form a compartment or analysis chamber. The energy may be selectively directed to the fluid device to form an analysis chamber at a predetermined location within the fluid device. Selective energy direction may be performed using a spatial energy modifier (e.g., a mask as described herein). In some embodiments, the method may further include the step of detecting the biological component using a detector. Information from the detected biological component (e.g., the location of the biological component in the fluid device) may be used to form or generate a spatial energy modifier. For example, the spatial energy modifier may inhibit or prevent the direction of energy to a location adjacent to the biological component. In some embodiments, the spatial energy modifier may selectively direct energy to a location adjacent to the biological component. In some embodiments, the spatial energy modifier may selectively direct energy onto the biological component.

[0291] In certain cases, the method may include the step of directing energy from an energy source to a predetermined part of a fluid device to form a polymer matrix structure at a predetermined location in the fluid device or in a predetermined pattern in the fluid device. In some embodiments, a detector (e.g., to detect the location of one or more biological components) may be used. In various embodiments, a detector may not be used. A certain number of polymer matrix structures may be formed or generated on or adjacent to biological components. In other words, there may be a sufficient number of biological components in the fluid device, and as a result, it is not necessary to first determine the location of the biological components before forming the polymer matrix structures.

[0292] The spatial energy modulating element may include a physical photomask, a virtual photomask, a physical electrode distribution pattern, a virtual electrode distribution pattern, a photolithography mask, a DMD system, or any other suitable mask. The physical or virtual photomask may, for example, prevent energy from being directed to a part of the fluid device while allowing energy to be directed to another part of the fluid device. The electrode distribution pattern may include electrically activating one or more electrodes (e.g., an array of electrodes) to allow energy to be directed to a part of the fluid device. The electrodes may be "off" where a polymer matrix does not need to be formed, or may be prevented from generating energy there. The electrodes may be "on" where a polymer matrix can be formed.

[0293] In some cases, the biological components may include cells or their transcriptomes. The cells may include eukaryotic cells, prokaryotic cells, fungal cells, algal cells, protists, plant cells, animal cells (e.g., human cells), or any other suitable cells. One or more reactions performed may include RNA sequencing. In some embodiments, one or more reactions performed may include transcriptome analysis (e.g., under different or varied conditions). For example, one or more reactions may be performed to analyze one or more effects on the transcriptome, such as temperature, small molecules, toxins, cell-cell interactions, or infection. In some embodiments, transcriptome analysis may provide gene expression profiles for cells or combinations of cells. In certain embodiments, one or more reactions performed may include readout based on hybridization of gene expression. The transcriptome may include messenger RNA (mRNA), long non-coding RNA (lncRNA), mitochondrial RNA, or total RNA. In some embodiments, other types of RNA (e.g., ribosomal RNA (rRNA)) may be analyzed and / or sequenced. RNA may contain at least 50 nucleotides (nt), 100 nt, 200 nt, 300 nt, 400 nt, 500 nt, 600 nt, 700 nt, 800 nt, 900 nt, 1,000 nt, 10,000 nt, or more. Biological components may contain any number of nucleotides between any two of the numbers referred to herein. RNA may be a double-stranded RNA (dsRNA) molecule or a single-stranded RNA (ssRNA) molecule.

[0294] Methods for analyzing two or more biological components are also provided herein. The method may include the step of introducing a first biological component and a second biological component into a fluid device. The method may include the step of forming a polymer matrix on or adjacent to the first biological component to form a first analysis chamber. The method may further include the step of forming a polymer matrix on or adjacent to the second biological component to form a second analysis chamber. The first analysis chamber may be adjacent to the second analysis chamber. The method may further include the step of analyzing one or more properties of the first biological component and / or the second biological component.

[0295] In some embodiments, one or more properties of the first biological component and / or the second biological component may include a response to an analyte, a response to a pharmaceutical agent, a response to an antimicrobial agent, production of a target compound by cells or a community of cells, production of a target molecule, production of nucleic acids, production of proteins, or any other suitable response. One or more properties may include at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 25, 50, 100, or more properties of the first biological component. One or more properties may include at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 25, 50, 100, or more properties of the second biological component. In some cases, properties of the first biological component may be compared to the same properties of the second biological component. In various embodiments, 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 25, 50, 100, or more properties of the first biological component and / or the second biological component may be analyzed or monitored. In certain embodiments, properties of the first biological component may be compared with different properties of the second biological component. For example, a first property of the first biological component may include a response to an analyte or a pharmaceutical agent in which the first biological component causes the production of a compound or molecule (e.g., an antibody, protein, enzyme, etc.). Then, a second property of the second biological component may include a response of the second biological component to the molecule or compound produced by the first biological component.

[0296] In some cases, the first and second properties of the first or second biological component may be analyzed. For example, the first and second properties may be analyzed in the first biological component in response to compounds or molecules produced by the second biological component. This may be used to elucidate one or more interactions between the first and second biological components.

[0297] The interaction may include a communication (e.g., a biological communication) between a first biological component and a second biological component. In some cases, the interaction may include a biochemical communication between a first biological component and a second biological component. In some embodiments, the biological communication may include a molecule containing a protein, nucleic acid, cytokine, chemokine, a combination thereof, or any other suitable molecule. The molecule may be generated by the first biological component or by the second biological component. In some cases, more than two biological components may be localized in analytical chambers formed adjacent to each other so that three or more properties of three or more biological components can be analyzed. In some embodiments, this may be used to investigate interactions between three or more biological components. In various embodiments, interactions between two, three, four, five, six, seven, ten, fifteen, twenty-five, fifty, one hundred, or more biological components may be investigated.

[0298] In some cases, the first and second biological components may include different cell types (e.g., a first cell type and a second cell type). In certain embodiments, the first and second biological components may include similar cell types, e.g., cell types having various genotypes or phenotypes. The response to the analyte or pharmaceutical compound (e.g., the level of response) can be compared between the two different cell types. In some cases, the amount of the target compound or the amount of molecular production between the two different cell types can be analyzed and / or compared. In some embodiments, the first and second biological components may remain localized within the first and second analysis chambers, respectively, during the analysis of their properties. Such analyses or experiments can be scaled up (e.g., in parallel) to evaluate or analyze 3, 4, 5, 6, 7, 8, 9, 10, 15, 25, 50, 100, or more biological components.

[0299] Methods for identifying nucleic acid molecules are also provided herein. These methods may include the capture of biological components. The biological components may then be encapsulated by a hydrogel, which may form an analysis chamber. In some embodiments, the nucleic acid molecules can be extracted and released within the analysis chamber. These nucleic acids may be sequenced by next-generation sequencing, which may be performed by short-read sequencing, nanopore sequencing, synthesis sequencing, in situ hybridization sequencing, or any optical readout, such as using a microscope. Other preferred methods of sequencing are also within the scope of this disclosure. The methods may further include the step of detecting nucleic acid molecules in the absence of nucleic acid amplification.

[0300] In some embodiments, biological components, including cells, may be introduced into a fluid device. A polymer matrix may then be formed on or adjacent to the biological components to form a compartment or analysis chamber. The analysis chamber may be formed by selectively directing energy from an energy source to a fluid device, as described herein. In some embodiments, the analysis chamber may be dismantled by degrading the polymer matrix "on demand." On-demand polymer matrix degradation may involve selectively directing energy to the analysis chamber using an enzyme for digesting or depolymerizing the polymer matrix or any other suitable method for degrading the polymer matrix.

[0301] Cells may be lysed and release biological components (e.g., DNA or RNA). In certain embodiments, the biological components are released from the cells upon interaction with the reagent. In some embodiments, the reagent is an organic or inorganic molecule. In some embodiments, the organic or inorganic molecule is a pharmaceutical compound or a surfactant. In some embodiments, the reagent is a protein. In some embodiments, the reagent is a DNA aptamer. In some embodiments, the reagent is a bead containing a biomolecule. In some embodiments, the reagent is a biological species. In some embodiments, the biological species is a virus or a cell.

[0302] In some embodiments, biological components are released from cells upon exposure to an energy source. In some embodiments, the energy source is UV light for lysing cells. In some embodiments, the energy source is visible light for lysing cells. In some embodiments, UV light is used to activate a photoactivatable surfactant and lyse cells. In some embodiments, visible light is used to activate a photoactivatable surfactant and lyse cells.

[0303] As described herein, the polymer matrix may have a pore size or mean pore size that does not allow nucleic acid molecules to pass through or traverse the polymer matrix. In some cases, the analysis chamber may be used for sequencing library preparation and / or nucleic acid sequencing. Sequencing may be next-generation sequencing. Sequencing may be short-read sequencing, nanopore sequencing, sequencing by synthesis, sequencing by in situ hybridization, or optical readout using, for example, a microscope. One or more nucleic acid molecules in the analysis chamber may undergo a nucleic acid sequencing reaction, for example, DNA sequencing or RNA sequencing. A nucleic acid library may be constructed or generated. The polymer matrix may allow the passage of reagents as described herein. Reagents may include primers, adapters, enzymes, and other reagents used for the nucleic acid sequencing reaction.

[0304] In certain embodiments, nucleic acids may include DNA molecules or RNA molecules. DNA may be at least 100 bp in length. In some embodiments, DNA may contain at least 50 bp, 100 bp, 200 bp, 300 bp, 400 bp, 500 bp, 600 bp, 700 bp, 800 bp, 900 bp, 1,000 bp, 10 kilobase pairs (kbp), 100 kbp, 1 megabase pair (Mbp), 100 Mbp, 1 gigabase pair (Gbp), 10 Gbp, 100 Gbp, or more base pairs. Biological components may include DNA molecules containing any number of base pairs between the numbers mentioned herein. For example, DNA may contain 50 bp to 1,000 bp, 300 bp to 10 kbp, or 1,000 bp to 10 Gbp. RNA may be dsRNA. dsRNA may contain at least 50 bp, 100 bp, 200 bp, 300 bp, 400 bp, 500 bp, 600 bp, 700 bp, 800 bp, 900 bp, 1,000 bp, 10 kbp, or 100 kbp. Biological components may contain dsRNA molecules containing any number of base pairs between the numbers mentioned herein. For example, dsRNA may contain 50 bp to 1,000 bp, 300 bp to 10 kbp, or 1,000 bp to 100 kbp. RNA may be ssRNA. ssRNA may contain 50 nt to 100,000 nt. ssRNA may consist of 50nt-100nt, 50nt-1,000nt, 50nt-10,000nt, 50nt-100,000nt, 100nt-1,000nt, 10nt-100nt, 10nt-1,000nt, or 100nt-1,000nt. In some cases, ssRNA may be less than 50 nucleotides long. ssRNA may also be longer than 100,000 nucleotides.

[0305] In some embodiments, sequencing library construction may include barcoding. Barcoding may be specific to each analysis chamber for associating nucleic acid molecules with the analysis chamber or biological components (e.g., cells). In some embodiments, polymerase chain reaction (PCR) may be used for sequencing library construction. In some cases, PCR may not be used for purposes other than sequencing library construction, as the nucleic acid material may be retained within the analysis chamber and not diluted or lost. In some embodiments, sequencing library construction may include adapter ligation. In some embodiments, all steps necessary for sequencing nucleic acid molecules may be performed while the biological components and / or nucleic acids are localized in the analysis chamber. In some embodiments, library preparation includes transposase-assisted tagmentation of nucleic acid molecules (e.g., by using transposons to cleave and label genomic DNA).

[0306] Methods for processing cells to determine the cellular transcriptome are also provided herein. In some embodiments, the method does not involve nucleic acid amplification. In other embodiments, the method may include nucleic acid amplification. The method may include the step of processing cells to determine the cellular epigenome. In some embodiments, the biological components, including cells, may be introduced into a fluid device provided herein. A polymer matrix may be formed on or adjacent to the biological components to form an analysis chamber (e.g., around or enclosing the biological components). The analysis chamber may at least partially or completely surround the cells. The analysis chamber may be formed by selectively directing energy from an energy source to a fluid device, as described herein. In some embodiments, the analysis chamber may be dismantled or removed by degrading the polymer matrix. The polymer matrix may be degraded "on demand," as provided herein.

[0307] Biological components or their products may be analyzed within the analysis chamber. In some cases, the biological components or their products may be eluted and transferred to another device for analysis. For example, nucleic acid material or protein product derived from a biological component (e.g., cells, bacteria, viruses, etc.) may be extracted and / or processed (e.g., tagged, barcoded, etc.) within the analysis chamber. The nucleic acid material or protein product may then be sequenced within the analysis chamber of a fluid device described herein. In some cases, the nucleic acid material or protein product may be eluted and transferred to another device (e.g., a sequencing flow cell) for sequencing (e.g., using a sequencing device).

[0308] The cells may include mammalian cells (e.g., human cells), fungal cells, bacterial cells, algal cells, protists, plant cells, tumor spheroids, or combinations thereof. In some embodiments, the cell's genome, transcriptome, proteome, epigenome, methylome, secretome, or metabolome may be extracted, for example, by lysing the cell. In some cases, one or more proteins produced and / or released by the cell can be analyzed without lysing the cell. In some cases, the cell's genome, transcriptome, proteome, epigenome, methylome, secretome, and / or metabolome can be studied, analyzed, and / or sequenced (where appropriate) while the cell and / or the cell's genome, transcriptome, proteome, epigenome, methylome, secretome, or metabolome may remain in the analysis chamber or substantially remain in the analysis chamber. In some embodiments, nucleic acids in the cell may remain in the analysis chamber. Therefore, nucleic acid amplification may be avoided or made unnecessary. In some cases, one or more cells may be processed in an analysis chamber to identify and study the genome, transcriptome, proteome, epigenome, methylome, secretome, and / or metabolome in one or more cells.

[0309] The first and / or second surfaces of the fluid device (similar to surfaces 101 and 102 in Figure 1) may contain a detection element. The detection element may contain one or more reagents or biochemical sensors. In some cases, reagents for studying the genome, transcriptome, proteome, epigenome, methylome, secretome, and / or metabolome of a cell may be introduced into the fluid device. The polymer matrix may contain pores that allow reagents to pass through, but the pores do not have to allow nucleic acids or proteins (e.g., cellular) to traverse the polymer matrix.

[0310] Methods for identifying multiple nucleic acid molecules from multiple cells without barcoding individual nucleic acid molecules of multiple nucleic acid molecules are also provided herein. The method may include the step of sequencing the multiple nucleic acid molecules. In some embodiments, the multiple biological components may be introduced into a fluid device. A polymer matrix may be formed on or adjacent to the biological components derived from the multiple biological components to form an analysis chamber that surrounds or encapsulates the biological components at least partially or completely. The analysis chamber may be formed by selectively directing energy from an energy source to a fluid device, as described herein. In some embodiments, the analysis chamber may be dismantled by decomposing the polymer matrix "on demand," as described herein. Multiple nucleic acid molecules of biological components (e.g., cells, bacteria, viruses, etc.) may be extracted in an analysis chamber formed surrounding the biological components. The nucleic acid molecules may then be sequenced within the analysis chamber. The sequence readout may be performed within the analysis chamber. Thus, the need for barcoding to associate the nucleic acid molecules and biological components can be avoided.

[0311] Multiple nucleic acid molecules may be extracted from multiple cells. In some cases, cellular nucleic acid molecules may be extracted from cells by lysing the cells in an analysis chamber (e.g., a compartment or chamber surrounding the cells). The analysis chamber may contain and / or localize the nucleic acids derived from the cells within the analysis chamber. Therefore, multiple nucleic acid molecules from multiple cells may be localized in individual and / or separate analysis chambers. In some embodiments, a sequencing process (e.g., nucleic acid library construction, sequencing, etc.) may be performed on multiple nucleic acid molecules that can be isolated or localized in separate analysis chambers. Therefore, barcoding to identify the cells that produce the nucleic acid molecules in multiple nucleic acid molecules may be avoided or made unnecessary.

[0312] Computer system This disclosure provides a computer system programmed to perform the methods of this disclosure. Figure 15 shows a computer system 1501 that may be programmed to perform the methods described herein, or may be otherwise configured. The computer system 1501 may control various aspects of this disclosure, such as, for example, the steps of identifying biological components, detecting barcodes, generating spatial modifiers (e.g., masks), supplying energy from an energy source, or detecting or measuring local parameters using sensors. The detector may be a camera (e.g., a fluorescence camera), such as a charge-coupled device (CCD) camera that can collect optical signals and positional information from multiple sources distributed across a planar area. The computer system 1501 may be a computer system that can be remotely installed on or with respect to a user's electronic device. The electronic device may be a mobile electronic device.

[0313] The computer system 1501 includes a central processing unit (CPU, also referred to herein as “processor” and “computer processor”) 1505, which may be a single-core or multi-core processor, or multiple processors for parallel processing. The computer system 1501 also includes memory or storage locations 1510 (e.g., random-access memory, read-only memory, flash memory), electronic storage devices 1515 (e.g., hard disks), communication interfaces 1520 for communicating with one or more other systems (e.g., network adapters), and peripheral devices 1525, e.g., caches, other memory, data storage, and / or electronic display adapters. The memory 1510, storage devices 1515, interfaces 1520, and peripheral devices 1525 communicate with the CPU 1505 through a communication bus (solid line), such as a motherboard. The storage device 1515 may be a data storage device (or data repository) for storing data. Computer system 1501 may be operationally connected to a computer network ("Network") 1530 with the help of a communication interface 1520. Network 1530 may be the Internet, the Internet and / or an extranet, or an intranet and / or extranet that can communicate with the Internet. In some cases, Network 1530 may be a telecommunications and / or data network. Network 1530 may include one or more computer servers, which may enable distributed computing such as cloud computing. In some cases, Network 1530 may, with the help of computer system 1501, run a peer-to-peer network, which may enable devices to connect to computer system 1501 to act as clients or servers.

[0314] The CPU 1505 can execute a sequence of machine-readable instructions that may be embodied in a program or software. Instructions may be stored in a memory location such as memory 1510. Instructions can instruct the CPU 1505 to subsequently be programmed or otherwise configured to perform the methods of this disclosure. Examples of operations performed by the CPU 1505 include fetching, decoding, executing, and writing back.

[0315] The CPU 1505 may be part of a circuit, such as an integrated circuit. One or more other components of system 1501 may be included in the circuit. In some cases, the circuit may be an application-specific integrated circuit (ASIC).

[0316] The storage device 1515 can store files such as drivers, libraries, and saved programs. The storage device 1515 can also store user data, such as user preferences and user programs. In some cases, the computer system 1501 may include one or more additional data storage devices outside of the computer system 1501, located on a remote server that can communicate with the computer system 1501, for example, via an intranet or the internet.

[0317] Computer system 1501 can communicate with one or more remote computer systems via network 1530. For example, computer system 1501 can communicate with a user's remote computer system (e.g., laptop, personal computer, tablet, or mobile phone). Examples of remote computer systems include personal computers (e.g., portable PCs), slate or tablet PCs (e.g., Apple® iPad®, Samsung® Galaxy Tab), telephones, smartphones (e.g., Apple® iPhone®, Android® compatible devices, Blackberry®), or personal digital assistants. The user can access computer system 1501 via network 1530.

[0318] The methods described herein can be implemented by machine-executable code (e.g., a computer processor) stored in an electronic storage location of the computer system 1501, such as memory 1510 or an electronic storage device 1515. The machine-executable code or machine-readable code may be provided in the form of software. During use, the code may be executed by the processor 1505. In some cases, the code may be retrieved from the storage device 1515 and stored in memory 1510, ready for access by the processor 1505. In some situations, the electronic storage device 1515 may be omitted, and the machine-executable instructions are stored in memory 1510.

[0319] The code may be pre-compiled and configured for use with a machine having a processor adapted to run the code, or it may be compiled during runtime. The code may be supplied in a programming language that can be chosen to allow the code to be run in the pre-compiled or compiled form.

[0320] Embodiments of systems and methods provided herein, for example, computer system 1501, can be embodied in programming. Various embodiments of technology can be considered “products” or “manufactured goods,” typically in the form of machine (or processor) executable code and / or related data that can be mounted on or embodied in some kind of machine-readable medium. Machine executable code can be stored in memory (e.g., read-only memory, random-access memory, flash memory) or electronic storage devices such as hard disks. “Storage” type media can include any or all of the tangible memory of a computer, processor, etc., or related modules thereof, for example, various semiconductor memories, tape drives, disk drives, etc., that can provide non-temporary storage at any time for software programming. All or part of the software may sometimes be communicated through the Internet or various other telecommunication networks. For example, such communication may enable the loading of software from one computer or processor to another, for example, from a management server or host computer to an application server computer platform. Therefore, other types of media that may have software elements include optical waves, radio waves, and electromagnetic waves, such as those used across physical interfaces between local devices, through wired and optical terrestrial network lines, and over various air links. Physical elements with such waves, e.g., wired or wireless links, optical links, etc., may also be considered media with software. As used herein, unless limited to non-temporary tangible “storage” media, terms such as computer or machine “readable media” refer to any medium involved in providing instructions to a processor for execution.

[0321] Therefore, machine-readable media, such as computer executable code, can take many forms, including, but are not limited to, tangible storage media, carrier media, or physical transmission media. Non-volatile storage media include, for example, optical disks or magnetic disks, such as any storage device in any computer, such as those that may be used to implement a database shown in a drawing. Volatile storage media include dynamic memory, such as the main memory of such a computer platform. Tangible transmission media include coaxial cables, copper wires, and optical fibers, and wires including buses in computer systems. Carrier media can take the form of electrical or electromagnetic signals, or acoustic or optical waves, such as those generated during radio wave (RF) and infrared (IR) data communications. Therefore, common forms of computer-readable media include, for example, floppy disks, flexible disks, hard disks, magnetic tapes, any other magnetic media, CD-ROMs, DVDs or DVD-ROMs, any other optical media, punched card paper tapes, any other physical storage media having a pattern of holes, RAM, ROMs, PROMs and EPROMs, FLASH®-EPROMs, any other memory chips or cartridges, carriers that transport data or instructions, cables or links that transport such carriers, or any other media from which a computer can read programming code and / or data. Many of these forms of computer-readable media can be involved in transporting one or more sequences of one or more instructions to a processor for execution.

[0322] The computer system 1501 may include, or communicate with, an electronic display 1535 that includes a user interface (UI) 1540 for providing, for example, images of biological components, barcodes, signals or measurements of local parameters. Examples of UIs include, but are not limited to, graphical user interfaces (GUIs) and web-based user interfaces.

[0323] The methods and systems of this disclosure can be implemented by one or more algorithms. The algorithms may be implemented by software when executed by the central processing unit 1505. The algorithms may, for example, identify biological components, detect barcodes, generate spatial modifiers (e.g., masks), supply energy from an energy source, and detect or measure local parameters using sensors.

[0324] Embodiment Embodiment 1. A system comprising a fluid device containing one or more biological components and one or more polymer precursors, and at least one energy source communicating with the fluid device, wherein the at least one energy source supplies energy to the fluid device to cause the one or more polymer precursors to form at least one polymer matrix on or adjacent to the biological components.

[0325] Embodiment 2. The system according to Embodiment 1, wherein the fluid device includes a channel through which it is disposed, the first surface being disposed along a portion of the channel, and the second surface being disposed opposite the first surface.

[0326] Embodiment 3. The system according to Embodiment 1 or Embodiment

[0325] , wherein the fluid device includes a chamber disposed therein, the first surface being disposed along a portion of the chamber, and the second surface being disposed opposite the first surface.

[0327] Embodiment 4. The system according to Embodiment

[0325] or Embodiment

[0326] , wherein the first surface is the bottom surface and the second surface is the top surface.

[0328] Embodiment 5. The system according to Embodiment 1, wherein the fluid device further includes one or more capture elements for immobilizing at least one of the one or more biological components at a location adjacent to the first surface that forms the immobilized biological components.

[0329] Embodiment 6. The system according to Embodiment 1, wherein the first surface is located adjacent to the energy source, and the energy source is an array of electrodes.

[0330] Embodiment 7. The system according to Embodiment

[0329] , wherein the electrode array supplies electrochemical energy to one or more polymer precursors to form an array of polymer matrices.

[0331] Embodiment 8. The system according to Embodiment 1, wherein at least two of the one or more polymer precursors are coupled to the first surface to form a pattern on the first surface.

[0332] Embodiment 9. The system according to Embodiment

[0331] , wherein at least one polymer matrix is ​​formed on or adjacent to the pattern.

[0333] Embodiment 10. The system according to Embodiment

[0328] , wherein at least one polymer matrix is ​​coupled to the first surface.

[0334] Embodiment 11. The system according to Embodiment

[0333] , wherein the at least one polymer matrix extends from the first surface to the second surface such that the at least one polymer matrix surrounds at least a portion of the immobilized biological component.

[0335] Embodiment 12. The system according to any one of Embodiments 1 to

[0334] , wherein the at least one energy source is in communication with the fluid device by at least one of optical communication, electrochemical communication, electromagnetic communication, thermal communication, or microwave communication.

[0336] Embodiment 13. The system according to any one of Embodiments 1 to

[0334] , wherein the at least one energy source includes a photogenerating device, a thermogenerating device, an electrochemical generating device, an electrode, or a microwave device.

[0337] Embodiment 14. The system according to any one of Embodiments 1 to

[0336] , further comprising a photolithography device or a digital micromirror device (DMD) configured to control the spatial distribution of the energy from the energy source.

[0338] Embodiment 15. The system according to any one of Embodiments

[0328] to

[0337] , wherein one or more of the capture elements include a physical trap, a geometric trap, a well, an electrochemical trap, a chemical affinity trap, one or more magnetic particles, an electrophoretic trap, a dielectrophoretic trap, or a combination thereof.

[0339] Embodiment 16. The system according to Embodiment

[0338] , wherein the chemical affinity trap comprises streptavidin, an antibody, or a combination thereof.

[0340] Embodiment 17. The system according to Embodiment

[0338] , wherein the physical trap includes a polymer matrix.

[0341] Embodiment 18. The system according to Embodiment

[0340] , wherein the polymer matrix comprises a hydrogel.

[0342] Embodiment 19. The system according to Embodiment

[0338] , wherein the electrochemical trap includes a gold electrode, a platinum electrode, or an indium tin oxide (ITO) electrode.

[0343] Embodiment 20. The system according to any one of embodiments

[0328] to

[0342] , wherein one or more of the capturing elements are arranged in a pattern on the first surface or the second surface.

[0344] Embodiment 21. The system according to any one of Embodiments

[0328] to

[0343] , wherein one or more capture elements include a well, the diameter of the well is 1 μm (micrometer) to 50 μm, and the depth of the well is 0.1 μm to 100 μm.

[0345] Embodiment 22. The system according to any one of Embodiments

[0328] to

[0344] , wherein the one or more biological components are a plurality of biological components, and the plurality of biological components are coupled to the one or more capture elements.

[0346] Embodiment 23. The system according to any one of Embodiments 1 to

[0345] , wherein the fluid device is a microfluidic device or a nanofluidic device.

[0347] Embodiment 24. The system according to any one of Embodiments 1 to

[0345] , wherein the fluid device is used for nucleic acid sequencing.

[0348] Embodiment 25. The system according to Embodiment

[0347] , wherein the nucleic acid sequencing includes next-generation sequencing, short-read sequencing, nanopore sequencing, synthesis sequencing, in situ hybridization sequencing, or optical readout.

[0349] Embodiment 26. The system according to any one of Embodiments 1 to

[0348] , wherein one or more of the above-mentioned biological components include cells, cell lysates, nucleic acids, microbiomes, proteins, cell mixtures, spatially linked biological components, or metabolites.

[0350] Embodiment 27. The system according to Embodiment

[0349] , wherein the cell mixture comprises a first cell type and a second cell type, and the first cell type is different from the second cell type.

[0351] Embodiment 28. The system according to Embodiment

[0349] , wherein the cells are animal cells, plant cells, fungal cells, or bacterial cells.

[0352] Embodiment 29. The system according to Embodiment

[0351] , wherein the animal cells are human cells.

[0353] Embodiment 30. The system according to any one of Embodiments 1 to

[0351] , wherein one or more of the above-mentioned biological components include a tumor spheroid or a spatially linked biological sample.

[0354] Embodiment 31. The system according to any one of Embodiments

[0349] to

[0353] , wherein the nucleic acid is DNA with 100 base pairs or more, or RNA with 50 base pairs or more.

[0355] Embodiment 32. The system according to any one of Embodiments

[0349] to

[0354] , wherein the cell lysate contains DNA of 50 bp (base pairs) to 100 Gbp (gigabase pairs) or RNA of 50 bp to 100 kbp (kilobase pairs).

[0356] Embodiment 33. The system according to any one of Embodiments 1 to

[0355] , wherein the at least one polymer matrix comprises a hydrogel.

[0357] Embodiment 34. The system according to any one of Embodiments 1 to

[0356] , wherein the fluid device further comprises one or more polymer precursors.

[0358] Embodiment 35. The system according to Embodiment

[0356] , wherein one or more of the polymer precursors include a hydrogel precursor.

[0359] Embodiment 36. The system according to any one of Embodiments

[0328] to

[0358] , wherein the at least one polymer matrix inhibits the passage of the immobilized biological components.

[0360] Embodiment 37. The system according to any one of Embodiments 1 to

[0359] , wherein the at least one polymer matrix forms a cylinder shell or polygonal shell, including an internal space and polymer matrix walls.

[0361] Embodiment 38. The system according to Embodiment

[0360] , wherein the internal space has an inner diameter of 1 μm to 500 μm.

[0362] Embodiment 39. The system according to Embodiment

[0360] or Embodiment

[0361] , wherein the polymer matrix wall has a thickness of at least 1 μm (micrometer).

[0363] Embodiment 40. The system according to Embodiment

[0362] , wherein at least one of the polymer matrix walls is a hydrogel wall.

[0364] Embodiment 41. The system according to any one of Embodiments 1 to

[0363] , wherein at least one polymer matrix is ​​decomposable.

[0365] Embodiment 42. The system according to Embodiment

[0364] , wherein the degradation of the at least one polymer matrix is ​​"on demand".

[0366] Embodiment 43. The system according to Embodiment

[0364] or Embodiment

[0365] , wherein the at least one polymer matrix is ​​decomposable by at least one of (i) contacting the at least one polymer matrix with a cleavage reagent, (ii) heating the at least one polymer matrix to at least 90°C, or (iii) exposing the at least one polymer matrix to a wavelength of light that cleaves a photocleavable crosslinking agent that crosslinks the polymers of the at least one polymer matrix.

[0367] Embodiment 44. The system according to Embodiment

[0366] , wherein the at least one polymer matrix comprises a hydrogel, and the cleavage reagent decomposes the hydrogel.

[0368] Embodiment 45. The system according to Embodiment

[0366] , wherein the cleavage reagent comprises a reducing agent, an oxidizing agent, an enzyme, a pH-based cleavage reagent, or a combination thereof.

[0369] Embodiment 46. The system according to Embodiment

[0366] , wherein the cleavage reagent comprises dithiothreitol (DTT), tris(2-carboxyethyl)phosphine (TCEP), tris(3-hydroxypropyl)phosphine (THP), or a combination thereof.

[0370] Embodiment 47. The system according to any one of Embodiments 1 to

[0369] , wherein the at least one polymer matrix allows the passage of a reagent.

[0371] Embodiment 48. The system according to any one of Embodiments 1 to

[0370] , wherein the at least one polymer matrix comprises pores, and the average size of the pores is adjusted by applying heat, applying electricity, applying light, or a combination thereof using a chemical reagent.

[0372] Embodiment 49. The system according to Embodiment

[0370] or Embodiment

[0371] , wherein the reagent comprises at least one of an enzyme, a chemical substance, an oligonucleotide, or a primer having a size of less than 50 base pairs.

[0373] Embodiment 50. The system according to any one of Embodiments

[0370] to

[0372] , wherein the reagent comprises lysozyme, proteinase K, random hexamer, polymerase, transposase, ligase, catalytic enzyme, deoxynucleotide triphosphate, buffer, cell culture medium, or divalent cation.

[0374] Embodiment 51. The system according to any one of Embodiments 1 to

[0373] , wherein the at least one polymer matrix includes pores that are sized to allow the diffusion of a reagent across the at least one polymer matrix, but are too small to allow DNA or RNA to traverse the pores.

[0375] Embodiment 52. The above at least one polymer matrix comprises a hydrogel, wherein the hydrogel is polyethylene glycol (PEG)-thiol, PEG-acrylate, acrylamide, N,N'-bis(acryloyl)cystamine, PEG, polypropylene oxide (PPO), polyacrylic acid, poly(hydroxyethyl methacrylate) (PHEMA), poly(methyl methacrylate) (PMMA), poly(N-isopropylacrylamide) (PNIPAAm), poly(lactic acid) (PLA), poly(lactic acid-co-glycolic acid) (PLGA), polycaprolactone (PCL), poly(vinyl sulfonic acid) (PVSA), poly(L-aspartic acid), poly(L-glutamic acid), polylysine, agar, A system according to any one of Embodiments 1 to

[0374] , comprising agarose, alginate, heparin, alginate sulfate, dextran sulfate, hyaluronan, pectin, carrageenan, gelatin, chitosan, cellulose, collagen, bisacrylamide, diacrylate, diallylamine, triallylamine, divinyl sulfone, diethylene glycol diallyl ether, ethylene glycol diacrylate, polymethylene glycol diacrylate, polyethylene glycol diacrylate, trimethylopropane trimethacrylate, ethoxylated trimethylol triacrylate, or ethoxylated pentaerythritol tetraacrylate, or a combination or mixture thereof.

[0376] Embodiment 53. The system according to Embodiment

[0375] , wherein the hydrogel comprises an enzymatically degradable hydrogel, PEG-thiol / PEG-acrylate, acrylamide / N,N'-bis(acryloyl)cystamine (BACy), or PEG / PPO.

[0377] Embodiment 54. The system according to any one of Embodiments

[0328] to

[0376] , wherein the first surface, the second surface, or both of them include one or more barcodes.

[0378] Embodiment 55. The system according to Embodiment

[0377] , wherein one or more barcodes include an identifier for identifying the source of one or more biological components.

[0379] Embodiment 56. The system according to Embodiment

[0378] , wherein one or more of the above barcodes include an oligonucleotide.

[0380] Embodiment 57. The system according to Embodiment

[0378] , wherein the supply source includes a sample from which the one or more biological components are collected.

[0381] Embodiment 58. The system according to Embodiment

[0378] , wherein the supply source includes a physiological or anatomical supply source from which the one or more biological components are collected.

[0382] Embodiment 59. The system according to Embodiment

[0381] , wherein the anatomical source includes the organ of interest.

[0383] Embodiment 60. The system according to Embodiment

[0382] , wherein the subject is a human.

[0384] Embodiment 61. The system according to any one of Embodiments

[0377] to

[0383] , wherein one or more barcodes are configured to bind to one or more biological components, or molecules made from one or more biological components.

[0385] Embodiment 62. The system according to any one of Embodiments

[0328] to

[0384] , wherein the first surface, the second surface, or both of the above, comprises one or more compounds configured to bind to one or more biological components.

[0386] Embodiment 63. The system according to any one of Embodiments

[0328] to

[0385] , wherein the first surface, the second surface, or both are functionalized with a surface polymer.

[0387] Embodiment 64. The system according to Embodiment

[0386] , wherein the surface polymer is functionalized with oligonucleotides.

[0388] Embodiment 65. The system according to Embodiment

[0386] , wherein the surface polymer is functionalized with an antibody, cytokine, chemokine, pro...

Claims

1. A method for processing biological samples, (a) A step of introducing the biological sample into a fluid device, wherein the biological sample comprises biological components, and the biological components comprise cells, (b) A step of introducing a polymer precursor into the fluid device, (c) the step of determining the location of the biological components within the fluid device, and (d) A step of selectively polymerizing the polymer precursor to generate a polymer matrix from the polymer precursor in the fluid device. The polymer matrix contains, and the polymer matrix at least partially encapsulates the biological components. The method described above is A step of releasing an analyte from the cells, wherein the analyte includes messenger ribonucleic acid (RNA), The steps include capturing the messenger RNA with one or more capture oligonucleotides located in a portion of the fluid device encapsulated by the polymer matrix, and The steps include: degrading the polymer matrix and loading a reverse transcriptase reagent into the fluid device to copy the captured messenger RNA and produce complementary deoxyribonucleic acid (DNA). Methods that further include the above.

2. A method for processing a biological sample, (a) A step of introducing the biological sample into a fluid device, wherein the biological sample comprises biological components, and the biological components comprise cells, (b) A step of introducing a polymer precursor into the fluid device, (c) the step of determining the location of the biological components within the fluid device, and (d) A step of selectively polymerizing the polymer precursor to generate a polymer matrix from the polymer precursor in the fluid device. The polymer matrix contains, and the polymer matrix at least partially encapsulates the biological components. The method described above is A step of releasing an analyte from the cells, wherein the analyte includes messenger ribonucleic acid (RNA), A step of capturing the messenger RNA with one or more capture oligonucleotides located in a portion of the fluid device encapsulated by the polymer matrix, The steps include: degrading the polymer matrix and loading a reverse transcriptase reagent into the fluid device to copy the captured messenger RNA and produce complementary deoxyribonucleic acid (DNA); The step of sequencing the complementary DNA. Methods that further include the above.

3. A method for processing a biological sample, (a) A step of introducing the biological sample into a fluid device, wherein the biological sample comprises biological components, and the biological components comprise cells, (b) A step of introducing a polymer precursor into the fluid device, (c) the step of determining the location of the biological components within the fluid device, and (d) A step of selectively polymerizing the polymer precursor to generate a polymer matrix from the polymer precursor in the fluid device. The polymer matrix contains, and the polymer matrix at least partially encapsulates the biological components. The method described above is A step of releasing an analyte from the cells, wherein the analyte includes messenger ribonucleic acid (RNA), and A step of capturing the messenger RNA with one or more capture oligonucleotides located in a portion of the fluid device encapsulated by the polymer matrix, wherein each of the one or more capture oligonucleotides includes a barcode indicating its position within the fluid device. Methods that further include the above.

4. A method for processing a biological sample, (a) A step of introducing the biological sample into a fluid device, wherein the biological sample contains biological components, (b) A step of introducing a polymer precursor into the fluid device, (c) the step of determining the location of the biological components within the fluid device, (d) A step of selectively polymerizing the polymer precursor to produce a polymer matrix from the polymer precursor in the fluid device, wherein the polymer matrix at least partially encapsulates the biological components, the polymer matrix comprises a hydrogel, and the hydrogel comprises polyacrylamide and poly(ethylene glycol). Methods that include...

5. A method for processing a biological sample, (a) A step of introducing the biological sample into a fluid device, wherein the biological sample contains biological components, (b) A step of introducing a polymer precursor into the fluid device, (c) the step of determining the location of the biological components within the fluid device, and (d) A step of selectively polymerizing the polymer precursor to produce a polymer matrix from the polymer precursor in the fluid device, wherein the polymer matrix comprises a polymer matrix that at least partially encapsulates the biological components and is degradable. Includes, The method described above is The step of degrading the polymer matrix with a degrading agent selected from the group consisting of dithiothreitol (DTT), tris(2-chloroethyl) phosphate (TCEP), tetrahydropyran (THP), and sodium periodate (NaIO4). Methods that further include the above.

6. A method for processing a biological sample, (a) A step of introducing the biological sample into a fluid device, wherein the biological sample contains biological components, (b) A step of introducing a polymer precursor into the fluid device, (c) the step of determining the location of the biological components within the fluid device, and (d) A step of selectively polymerizing the polymer precursor to produce a polymer matrix from the polymer precursor in the fluid device, wherein the polymer matrix includes a polymer matrix that at least partially encapsulates the biological components and is photodegradable. Includes, The method described above is The step of photodegrading the polymer matrix by selectively projecting a light beam onto the polymer matrix. Methods that further include the above.

7. The method according to claim 3, wherein the polymer matrix forms a chamber, and the inside of a portion of the chamber contains the biological components.

8. The method according to claim 3, wherein the polymer matrix comprises a hydrogel.

9. The method according to claim 3, wherein the fluid device includes a surface having one or more trapping elements configured to couple to the biological components.

10. The method according to claim 9, wherein the one or more capture elements include one or more functional groups that can interact with the biological components.

11. The method according to claim 10, wherein one or more functional groups include a complementary DNA sequence that targets the DNA or RNA of the biological component.

12. The method according to claim 10, wherein the one or more functional groups include fibronectin.

13. The method according to claim 10, wherein one or more of the functional groups include an arginylglycylaspartate (RGD) peptide.

14. The method according to claim 10, wherein the one or more functional groups include an antibody.

15. The method according to claim 3, wherein the step of determining the location of the biological components within the fluid device in (c) includes imaging the fluid device using a detector.

16. The method according to claim 15, wherein the detector is coupled to an energy source configured to emit energy.

17. The method according to claim 16, wherein the energy source is a photogenerating device and the energy includes light.

18. The method according to claim 17, wherein the energy source is in optical communication with the fluid device.

19. The method of claim 18, further comprising the step of generating a virtual mask based on the location of the biological components in the fluid device determined in (c).

20. The method according to claim 19, further comprising the step of projecting the virtual mask using the light emitted from the photogenerating device.

21. The method according to claim 19, wherein the light projected from the virtual mask is used in (d) to selectively polymerize the polymer precursor to produce the polymer matrix.

22. The method according to claim 19, wherein the virtual mask is generated from a spatial light modulator (SLM).

23. The method according to claim 22, wherein the SLM is a digital micromirror device (DMD).

24. The method according to claim 5, wherein the biological component includes cells.

25. The method according to claim 24, further comprising the step of releasing an analyte from the cells.

26. The method according to claim 25, wherein the analyte comprises messenger ribonucleic acid (RNA), and the method further comprises the step of capturing the messenger RNA with one or more capture oligonucleotides located in a portion of the fluid device encapsulated by the polymer matrix.

27. The method according to claim 26, further comprising the steps of degrading the polymer matrix and loading a reverse transcriptase reagent into the fluid device to copy the captured messenger RNA and produce complementary deoxyribonucleic acid (DNA).

28. The method according to claim 27, further comprising the step of sequencing the complementary DNA.

29. The method according to claim 26, wherein each of the one or more captured oligonucleotides includes a barcode indicating its position within the fluid device.

30. The method according to claim 3, wherein the polymer matrix includes polymer matrix walls having an annular cross-section.

31. The method according to claim 3, further comprising the step of performing one or more functional assays on the biological components, wherein the one or more functional assays evaluate cell viability, cell morphology, cell secretion, cell response, cell-cell interactions, or any combination thereof.

32. The method according to claim 31, wherein the one or more functional assays are performed using bright-field phase-contrast imaging or fluorescence imaging of the biological components.

33. The method according to claim 3, further comprising the step of selecting the biological components from the biological sample based on one or more optical signals of the biological components, prior to (d).

34. The method according to claim 3, wherein the fluid device includes a channel having an inlet and an outlet, the channel includes a first surface and a second surface, the second surface is positioned opposite the first surface, the polymer matrix forms a wall extending between the first surface and the second surface, thereby forming a chamber, the chamber encapsulates the biological components.

35. The method according to claim 3, wherein the polymer matrix comprises a hydrogel, and the hydrogel comprises polyacrylamide and poly(ethylene glycol).

36. The polymer matrix comprises a degradable polymer matrix, and the method comprises dithiothreitol (DTT), tris(2-chloroethyl) phosphate (TCEP), tetrahydropyran (THP), and sodium periodate (NaIO). ４ The method according to claim 3, further comprising the step of degrading the polymer matrix with a degrading agent selected from the group consisting of ).

37. The method according to claim 3, wherein the polymer matrix comprises a photodegradable polymer matrix, and the method further comprises the step of photodegrading the polymer matrix by selectively projecting a light beam onto the polymer matrix.

38. The method according to claim 3, wherein the introduction step in (a) and the introduction step in (b) are performed simultaneously.

39. The method according to claim 5, wherein the introduction step in (a) and the introduction step in (b) are performed simultaneously.

40. The method according to claim 5, wherein the polymer matrix includes polymer matrix walls having an annular cross-section.

41. The method of claim 5, wherein the step of determining the location of the biological components in the fluid device in (c) includes imaging the fluid device using a detector.

42. The method according to claim 5, wherein the fluid device includes a channel having an inlet and an outlet, the channel including a first surface and a second surface, the second surface being positioned opposite the first surface, the polymer matrix forming a wall extending between the first surface and the second surface, thereby forming a chamber, the chamber encapsulating the biological components.

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