Method of screening single cells for the production of biologically active agents
First Claim
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1. A method of obtaining protein and nucleic acid sequence information about proteins produced from isolated cells of interest, comprising the steps of:
- placing a plurality of individual cells in a plurality of individual wells, wherein each cell produces at least one protein of interest;
contacting said at least one protein of interest in the wells with a first surface comprising at least one protein binding agent, which selectively binds at least one of the proteins of interest and which first surface comprises addressable regions that are relatable to the individual wells;
determining one or more properties of said one or more proteins of interest and relating the one or more properties to a particular addressed region of the first surface thereby identifying a well of particular interest;
lysing the cells in the wells;
hybridizing nucleic acids from the lysed cells with oligonucleotide probes comprising tags bound to a second surface, wherein the second surface comprises regions relatable to the individual wells and wherein the tags in each region are different;
converting the nucleic acids into copy nucleic acids that comprise the tags specific to the regions on the second surface;
pooling the tagged copy nucleic acids;
sequencing the tagged copy nucleic acids; and
utilizing the tags to associate the copy nucleic acid sequences to a particular well, which associates the copy nucleic acid sequences with properties of the proteins of interest.
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Abstract
This invention generally relates to a methods, devices and kits for screening single cells for the production of one or more biologically active agents of interest, such as a protein, nucleic acid, or a protein and the nucleic acid encoding same.
100 Citations
18 Claims
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1. A method of obtaining protein and nucleic acid sequence information about proteins produced from isolated cells of interest, comprising the steps of:
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placing a plurality of individual cells in a plurality of individual wells, wherein each cell produces at least one protein of interest; contacting said at least one protein of interest in the wells with a first surface comprising at least one protein binding agent, which selectively binds at least one of the proteins of interest and which first surface comprises addressable regions that are relatable to the individual wells; determining one or more properties of said one or more proteins of interest and relating the one or more properties to a particular addressed region of the first surface thereby identifying a well of particular interest; lysing the cells in the wells; hybridizing nucleic acids from the lysed cells with oligonucleotide probes comprising tags bound to a second surface, wherein the second surface comprises regions relatable to the individual wells and wherein the tags in each region are different; converting the nucleic acids into copy nucleic acids that comprise the tags specific to the regions on the second surface; pooling the tagged copy nucleic acids; sequencing the tagged copy nucleic acids; and utilizing the tags to associate the copy nucleic acid sequences to a particular well, which associates the copy nucleic acid sequences with properties of the proteins of interest. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17)
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18. A method comprising:
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placing a plurality of cells in a plurality of wells; allowing the cells in the wells to exist under conditions such that the cells produce antibodies; contacting the antibodies in the wells with a first surface comprising regions relatable to the wells, wherein the first surface comprises an antibody binding agent; contacting the antibodies bound to the regions on the first surface with one or more antibody binding agents;
determining binding information related to binding of the one or more antibody binding agents to the antibodies on the regions of the first surface;ysing the cells in the wells;
incorporating a tag in nucleic acid copies of the nucleic acids from the lysed cells wherein each tag is uniquely associated with a single well;pooling nucleic acid copies from the plurality of wells; sequencing the pooled nucleic acids using ultra high throughput DNA sequencing technology; and using the tags to associate nucleotide sequences with wells, which associates nucleotide sequences with binding information from the antibodies.
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Specification