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US 11,926,863 B1Granted
Solid state single cell method for analyzing fixed biological cells
Assignee10x Genomics, Inc.
InventorStephane Claude Boutet
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Abstract
A method of analyzing single biological particles, such as cells or nuclei, that maintains the single biological particles in a state of relative isolation during decrosslinking and subsequent processing is provided. In the case of cells, the method prevents cellular analytes from each individual cell from leaving the cell site and diffusing toward adjacent cells, while permitting transmission of a decrosslinking agent, followed by processing of the cellular analytes from the individual cells by barcoding and/or imaging of the cellular analytes.
Claims19
A method of analysis for aldehyde fixed cells comprising:
- providing a substrate comprising a plurality of capture regions, wherein a capture region of the plurality of capture regions comprises:
- a plurality of antibodies capture moiety and a plurality of barcode molecules, wherein a barcode molecule of the plurality of barcode molecules comprises a common barcode sequence and an analyte a nucleic acid binding sequence;
- contacting a plurality of aldehyde fixed cells with the substrate, wherein an aldehyde fixed cell of the plurality of aldehyde fixed cells binds to an antibody of the plurality of antibodies, thereby generating a captured aldehyde fixed cell at the capture region;
- applying a polydimethyl siloxane permeable coating to the substrate, thereby immobilizing the captured aldehyde fixed cell;
- reversing fixation of the aldehyde captured fixed cell at the capture region to release a plurality of nucleic acids from the captured aldehyde fixed cell and hybridizing a nucleic acid of the plurality of nucleic acids to the nucleic acid binding sequence of the barcode molecule;
- and
- generating a plurality of barcoded molecules from the plurality of nucleic acids and the plurality of barcode molecules, wherein a barcoded molecule of the plurality of barcoded molecules comprises the common barcode sequence, or a complement thereof, and a sequence corresponding to the nucleic acid, or a complement thereof.
4 more elements
Dependencies are inferred from the claim text.