Methods of detecting analytes
AssigneeSpatial Transcriptomics AB
InventorsJonas Frisen, Patrik Stahl, Joakim Lundeberg
Life of this patent
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Abstract
Localized detection of RNA in a tissue sample that includes cells is accomplished on an array. The array include a number of features on a substrate. Each feature includes a different capture probe immobilized such that the capture probe has a free 3′ end. Each feature occupies a distinct position on the array and has an area of less than about 1 mm2. Each capture probe is a nucleic acid molecule, which includes a positional domain including a nucleotide sequence unique to a particular feature, and a capture domain including a nucleotide sequence complementary to the RNA to be detected. The capture domain can be at a position 3′ of the positional domain.
Claims24
A method for localized detection of RNA in a tissue section, the method comprising:
- (a) providing an array comprising a plurality of features on a substrate, wherein each feature occupies a distinct position on the array, wherein a feature of the plurality of features comprises a plurality of capture probes immobilized thereon, and wherein the plurality of capture probes comprise a nucleic acid molecule having the following domains oriented 5′ to 3′:
- (i) a first sequence comprising a positional domain comprising a nucleotide sequence unique to the feature;
- and
- (ii) a second sequence comprising a capture domain comprising a nucleotide sequence complementary to the RNA to be detected;
- (b) contacting the array with the tissue section and allowing at least one RNA of the tissue section to hybridize to the capture domain of the capture probe;
- (c) generating a cDNA molecule, by extending the capture probe using the hybridized RNA as an extension template, such that the generated cDNA molecule comprises the nucleotide sequence of the positional domain;
- (d) releasing the generated cDNA molecule, or a portion thereof, or a second strand complementary to the generated cDNA molecule, or a portion thereof, from the array, and
- (e) identifying the nucleotide sequence of the positional domain in the released cDNA molecule, or the complement thereof;
- and
- (f) correlating the nucleotide sequence of the positional domain with the distinct position on the array, thereby detecting the RNA in the tissue section.
Dependencies are inferred from the claim text.