Granted Patent
B2
US 12,529,093 · App. 18/769,778 · Granted Jan 20, 2026
Detecting tumor infiltrating cells
Assignee:
Singular Genomics Systems, Inc.
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Abstract
Disclosed herein, inter alia, are compositions and methods of use thereof for interrogating a tissue sample.
Claims (33)
1 . A method of detecting an immune cell in a tissue, said method comprising:
contacting the tissue with a polynucleotide probe and hybridizing a first hybridization sequence of the polynucleotide probe to a first sequence of an RNA molecule, and hybridizing a second hybridization sequence of the polynucleotide probe to a second sequence of the RNA molecule, wherein said RNA molecule is in the immune cell and comprises a target sequence between the first sequence and the second sequence;
extending the polynucleotide probe along the target sequence to generate a complement of the target sequence, and ligating the complement of the target sequence to the polynucleotide probe thereby forming a circular oligonucleotide;
extending a first oligonucleotide hybridized to the circular oligonucleotide with a polymerase to generate an extension product comprising the target sequence; and
hybridizing a second oligonucleotide to the extension product and sequencing the target sequence in the tissue, thereby detecting the immune cell.
2 . The method of claim 1 , wherein the immune cell is a T cell.
3 . The method of claim 2 , wherein the RNA molecule encodes for a chimeric antigen receptor.
4 . The method of claim 1 , wherein the RNA molecule comprises a T cell receptor alpha variable gene sequence (TRAV gene), T cell receptor alpha joining gene sequence (TRAJ gene), T cell receptor alpha constant gene sequence (TRAC gene), T cell receptor beta variable gene sequence (TRBV gene), T cell receptor beta diversity gene sequence (TRBD gene), T cell receptor beta joining gene sequence (TRBJ gene), T cell receptor beta constant gene sequence (TRBC gene), T cell receptor gamma variable gene sequence (TRGV gene), T cell receptor gamma joining gene sequence (TRGJ gene), T cell receptor gamma constant gene sequence (TRGC gene), T cell receptor delta variable gene sequence (TRDV gene), T cell receptor delta diversity gene sequence (TRDD gene), T cell receptor delta joining gene sequence (TRDJ gene), or T cell receptor delta constant gene sequence (TRDC gene).
5 . The method of claim 1 , wherein the immune cell is a NK cell.
6 . The method of claim 1 , wherein the immune cell is an allogenic cell.
7 . The method of claim 1 , wherein the immune cell is a non-allogenic cell.
8 . The method of claim 1 , further comprising detecting a population of immune cells in the tissue, wherein the immune cells of the population comprise the same target sequence.
9 . The method of claim 1 , further comprising detecting a plurality of populations of immune cells, wherein the immune cells of each population comprise the same target sequence.
10 . The method of claim 1 , wherein the polynucleotide probe comprises DNA, RNA, LNA, or a combination thereof.
11 . The method of claim 1 , wherein the polynucleotide probe consists of DNA.
12 . The method of claim 1 , wherein the tissue is immobilized to a solid support.
13 . The method of claim 1 , wherein the polynucleotide probe is about 50 to about 100 nucleotides.
14 . The method of claim 1 , wherein the polynucleotide probe is a single-stranded polynucleotide comprising an amplification primer binding sequence, a sequencing primer binding sequence, or both an amplification primer binding sequence and a sequencing primer binding sequence.
15 . The method of claim 1 , wherein the sequencing comprises sequencing-by-synthesis.
16 . The method of claim 1 , wherein sequencing comprises repeated cycles of labeled oligonucleotide hybridization and detection.
17 . The method of claim 1 , wherein sequencing comprises extending the second oligonucleotide by incorporating a labeled nucleotide, or labeled nucleotide analogue, and detecting the label for each incorporated nucleotide or nucleotide analogue.
18 . The method of claim 1 , wherein the target sequence is about 100 to about 200 nucleotides.
19 . The method of claim 1 , wherein the tissue comprises breast tissue, lung tissue, colon tissue, lymph tissue, kidney tissue, bone tissue, tonsil tissue, or brain tissue.
20 . The method of claim 1 , wherein the tissue comprises kidney tissue.
21 . The method of claim 1 , wherein the tissue comprises colon tissue.
22 . The method of claim 1 , wherein the tissue comprises brain tissue.
23 . The method of claim 1 , wherein the tissue comprises lung tissue.
24 . The method of claim 1 , wherein the tissue comprises breast tissue.
25 . The method of claim 1 , wherein the tissue comprises bone tissue.
26 . The method of claim 1 , wherein the tissue comprises lymph tissue.
27 . The method of claim 1 , further comprising detecting a protein in the tissue.
28 . The method of claim 27 , comprising detecting the protein in or on the immune cell.
29 . The method of claim 1 , further comprising contacting the tissue with a stain and detecting the stain, wherein the stain comprises an ER stain, Golgi stain, F-actin stain, lysosomal stain, mitochondrial stain, nucleolar stain, or plasma membrane stain.
Assignments (2)
SECURITY INTEREST
Recorded Mar 7, 2025
From: SINGULAR GENOMICS SYSTEMS, INC.
To: FIRST-CITIZENS BANK & TRUST COMPANY
Reel/Frame 070440/0465 →
ASSIGNMENT OF ASSIGNOR'S INTEREST
Recorded Aug 22, 2024
From: GLEZER, ELI N.
To: SINGULAR GENOMICS SYSTEMS, INC.
Reel/Frame 068371/0252 →
Continuity (10)
Provisional Application
63209903
· Jun 11, 2021
Provisional Application
63140703
· Jan 22, 2021
Provisional Application
63062047
· Aug 6, 2020
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