US 11,873,482 B2
Methods, compositions, and systems for spatial analysis of analytes in a biological sample
Hanyoup Kim, Pleasanton, CA (US); David Sukovich, Pleasanton, CA (US); Layla Katiraee, Pleasanton, CA (US); Augusto Manuel Tentori, Pleasanton, CA (US); Lauren Gutgesell, Pleasanton, CA (US); Janine Hensel, Pleasanton, CA (US); and Seayar Mohabbat, Pleasanton, CA (US)
Assigned to 10x Genomics, Inc., Pleasanton, CA (US)
Filed by 10x Genomics, Inc., Pleasanton, CA (US)
Filed on Apr. 27, 2023, as Appl. No. 18/140,312.
Application 18/140,312 is a continuation of application No. 17/976,422, filed on Oct. 28, 2022, granted, now 11,680,260.
Application 17/976,422 is a continuation in part of application No. 17/871,525, filed on Jul. 22, 2022, granted, now 11,618,897, issued on Apr. 4, 2023.
Application 17/871,525 is a continuation of application No. PCT/US2021/061401, filed on Dec. 1, 2021.
Claims priority of provisional application 63/252,323, filed on Oct. 5, 2021.
Claims priority of provisional application 63/148,825, filed on Feb. 12, 2021.
Claims priority of provisional application 63/128,796, filed on Dec. 21, 2020.
Prior Publication US 2023/0332138 A1, Oct. 19, 2023
This patent is subject to a terminal disclaimer.
Int. Cl. C12N 15/10 (2006.01)
CPC C12N 15/1065 (2013.01) 30 Claims
 
1. A method for processing a nucleic acid analyte in a biological sample mounted on a first substrate, the method comprising:
(a) providing the biological sample, wherein the biological sample is a tissue sample that was previously treated with a fixative comprising an aldehyde and embedded in an optimal cutting temperature (OCT) compound;
(b) hybridizing a first probe and a second probe to the nucleic acid analyte, wherein the first probe and the second probe each comprise a sequence that is substantially complementary to sequences of the nucleic acid analyte, and wherein the second probe comprises a capture probe binding domain;
(c) coupling the first probe and the second probe, thereby generating a connected probe;
(d) aligning the first substrate with a second substrate comprising an array, such that at least a portion of the biological sample is aligned with at least a portion of the array, wherein the array comprises a plurality of capture probes, wherein a capture probe of the plurality of capture probes comprises: (i) a spatial barcode and (ii) a capture domain;
(e) releasing the connected probe from the nucleic acid analyte when at least a portion of the biological sample is aligned with at least a portion of the array; and
(f) hybridizing the connected probe to the capture domain of the capture probe.