Patent ID: 11965208
Assignee: BECTON, DICKINSON AND COMPANY
Field: Biotechnology (Chemistry)
Classification: CPC C | IPC C

Claim 21:
22. A method of generating a spatial index of a plurality of cells associated with cell indexing oligonucleotides, comprising:
generating a plurality of cells associated with unique cell indexing oligonucleotides by n rounds of split-pool synthesis, wherein n is an integer greater than 1, wherein each cell is associated with a unique cell indexing oligonucleotide via a cellular component-binding reagent, wherein each cell indexing oligonucleotide comprises:
a cell indexing sequence comprising n cell indexing subsequences; and
a target sequence comprising a sequence complementary to a capture sequence of a stochastic label configured to hybridize to the target sequence, wherein the stochastic label comprises a target binding region comprising the capture sequence, wherein the target binding region is configured to hybridize to a nucleic acid target of a cell of the plurality of cells,

wherein n is greater than 1, and wherein each round of split-pool synthesis provides a cell indexing subsequence of the n cell indexing subsequences,
wherein the cell indexing sequence of a cell indexing oligonucleotide associated with a cell of the plurality of cells is different from the cell indexing sequence of cell indexing oligonucleotides associated with different cells of the plurality of cells;

partitioning the plurality of cells associated with the unique cell indexing oligonucleotides to a plurality of partitions on a substrate, wherein each partition of the plurality of partitions is a distinct spatial location on the substrate and comprises at most a single cell from the plurality of cells associated with the unique cell indexing oligonucleotides; and
generating a spatial index of the unique cell indexing oligonucleotides in the plurality of partitions using n pluralities of probes by sequentially hybridizing and imaging the n pluralities of probes annealing to the n cell indexing subsequences, wherein the spatial index identifies a distinct spatial location of the plurality of partitions on the substrate.