Methods and compositions related to microsphere surface gelation
The present disclosure relates generally to compositions and methods for generating hydrogel coatings on features. In some embodiments, hydrogel-coated features are prepared by horseradish peroxidase-mediated gelation of microspheres. Some embodiments relate to methods for generating hydrogel coatings using particle templated emulsification. The disclosure also relates to the use of hydrogel-coated features in arrays for spatial analysis of biological analytes in biological samples.
1 . A hydrogel-coated feature comprising:
(a) a solid particle; and
(b) a hydrogel encapsulating the solid particle, wherein the hydrogel comprises a crosslinked polymer and a plurality of moieties.
2 . The hydrogel-coated feature of claim 1 , wherein each moiety of the plurality of moieties comprises an oligonucleotide, wherein the oligonucleotide is configured to hybridize to a capture probe.
3 . The hydrogel-coated feature of claim 1 , wherein each moiety of the plurality of moieties comprises an organic functional group, wherein the organic functional group is capable of attachment to a capture probe.
4 . The hydrogel-coated feature of claim 3 , wherein the organic functional group is an amine group.
5 . The hydrogel-coated feature of claim 1 , wherein the hydrogel comprises an acrylamide copolymer.
6 . The hydrogel-coated feature of claim 5 , wherein the acrylamide copolymer is prepared from (i) acrylamide and 3-aminopropyl methacrylamide monomers or (ii) acrylamide, 3-aminopropyl methacrylamide, and 5′ acrydite oligonucleotide monomers.
7 . The hydrogel-coated feature of claim 6 , wherein the acrylamide copolymer is prepared (i) in the presence of sodium formate, (ii) in the presence of a polymerization initiator or (iii) in the presence of sodium formate and a polymerization initiator.
8 . The hydrogel-coated feature of claim 3 , wherein the organic functional group is phenol.
9 . The hydrogel-coated feature of claim 1 , wherein the solid particle is a microsphere, optionally wherein the microsphere has a diameter of about 1 micrometer to about 100 micrometers.
10 . The hydrogel-coated feature of claim 1 , wherein the hydrogel-coated feature comprises a plurality of capture probes and wherein each capture probe is attached to a moiety of the plurality of moieties within the hydrogel.
11 . The hydrogel-coated feature of claim 10 , wherein the capture probe comprises a barcode sequence and a capture domain.
12 . The hydrogel-coated feature of claim 11 , wherein the capture domain comprises a poly(T) sequence, a random sequence, a gene specific sequence, or a degenerate sequence.
13 . The hydrogel-coated feature of claim 10 , wherein the capture probe further comprises a unique molecular identifier (UMI) and/or a functional domain.
14 . The hydrogel-coated feature of claim 10 , wherein the feature comprises the plurality of capture probes at an average density of about 1,000 to about 100,000 capture probes per micrometer 2 .
15 . A substrate comprising a plurality of the hydrogel-coated features of claim 1 , wherein the hydrogel-coated features are immobilized on the substrate.
16 . The substrate of claim 15 , wherein the hydrogel-coated features are spatially ordered in an array and wherein each hydrogel-coated feature of the plurality of the hydrogel-coated features is associated with a unique spatial location on the array.
17 . The substrate of claim 15 , wherein the substrate comprises glass, silicon dioxide or a silicon wafer.
18 . A method for spatial analysis of a biological analyte in a biological sample, comprising:
(a) providing a substrate comprising hydrogel-coated features of claim 1 ;
(b) contacting the substrate with the biological sample under conditions sufficient for at least one biological analyte of the biological sample to hybridize to a capture probe on at least one hydrogel-coated feature; and
(c) determining (i) all or a part of the sequence of the biological analyte hybridized to the capture probe, or a complement thereof, and (ii) the spatial barcode, or a complement thereof, and using the determined sequence of (i) and (ii) to identify the location of the biological analyte in the biological sample.
19 . A method of preparing a hydrogel-coated feature, comprising:
(a) providing a solid particle; and
(b) encapsulating the solid particle with a hydrogel comprising a crosslinked polymer and a plurality of a moieties.
20 . A method for preparing a plurality of hydrogel-coated features comprising:
(a) suspending a plurality of features in an aqueous solution comprising a linear polymer, a surfactant, and an enzyme, thereby forming a suspension comprising the plurality of features;
(b) adding an oil to the suspension and emulsifying the suspension into a plurality of droplets, wherein a droplet of the plurality of droplets comprises a feature of the plurality of features coated with the linear polymer and the enzyme;
(c) contacting the plurality of droplets with a substrate for the enzyme, wherein the enzyme mediates crosslinking of the linear polymer to form a hydrogel on a feature of the plurality of features, thereby forming a plurality of hydrogel-coated features.
21 . A method of immobilizing a capture probe on a feature, comprising:
(a) encapsulating a solid particle with a hydrogel comprising a crosslinked polymer, and
(b) attaching the capture probe to a moiety within the hydrogel.
22 . A method for preparing a plurality of hydrogel-coated features comprising:
(a) suspending a plurality of features in an aqueous solution comprising a polymerizable monomer and a surfactant, thereby forming a suspension comprising the plurality of features;
(b) adding a partitioning oil to the suspension and emulsifying the suspension into a plurality of droplets, wherein a droplet of the plurality of droplets comprises a feature of the plurality of features coated with the monomer;
(c) contacting the plurality of droplets with a polymerization agent such that the monomer coating a feature of the plurality of features is polymerized to form a plurality of hydrogel-coated features.