Provided herein are methods for simultaneous spatio-temporal measurement of gene expression and cellular activity.

Provided herein are methods for simultaneous spatio-temporal measurement of gene expression and cellular activity.

Methods for determining a location of a feature on a spatial array include (a) providing an array of features on a substrate, where a feature of the array includes a barcoded oligonucleotide having, in a 5′ to 3′ direction, a spatial barcode, a cleavage domain, and a constant sequence; (b) hybridizing a priming oligonucleotide to the constant sequence; (c) extending the priming oligonucleotide using the barcoded oligonucleotide as a template; and (d) determining all or a portion of a sequence of the extended priming oligonucleotide corresponding to the spatial barcode, or a complement thereof, and a location of the extended priming oligonucleotide, and using the location of the extended priming oligonucleotide to determine the location of the feature on the spatial array.

A sample holder includes a first member featuring a first retaining mechanism configured to retain a first substrate that includes a sample, a second member featuring a second retaining mechanism configured to retain a second substrate that includes a reagent medium, and an alignment mechanism connected to at least one of the first and second members, and configured to align the first and second members such that the sample contacts at least a portion of the reagent medium when the first and second members are aligned.

This disclosure provides methods for spatial profiling of biological analytes present in a biological sample. Methods include generating feature arrays using a master/copy format using recessed arrays, and methods for using such arrays. For example spatially-tagged analyte capture analytes can be used in spatial detection in methods to determine the location of analytes (e.g., proteins) in biological samples.

Disclosed herein include systems, methods, compositions, and kits for performing intracellular AbSeq assays. There are provided, in some embodiments, methods for measuring intracellular target expression. The method can comprise fixing and permeabilizing a plurality of cells before contacting with a plurality of intracellular target-binding reagents capable of specifically binding to an intracellular target. Intracellular target-binding reagents can comprise an intracellular target-binding reagent specific oligonucleotide comprising a unique intracellular target identifier for the intracellular target-binding reagent specific oligonucleotide. The method can further comprise removing the permeabilizing agent and reversing the fixation.

Disclosed herein include systems, methods, compositions, and kits for performing intracellular AbSeq assays. There are provided, in some embodiments, methods for measuring intracellular target expression. The method can comprise fixing and permeabilizing a plurality of cells before contacting with a plurality of intracellular target-binding reagents capable of specifically binding to an intracellular target. Intracellular target-binding reagents can comprise an intracellular target-binding reagent specific oligonucleotide comprising a unique intracellular target identifier for the intracellular target-binding reagent specific oligonucleotide. The method can further comprise removing the permeabilizing agent and reversing the fixation.

Provided herein, among other things, are various compositions and methods for analyzing chromatin. In some embodiments, the composition may comprise a mixture of a nicking enzyme, four dNTPs, at least one labeled dNTP and, optionally, a polymerase. In some embodiments, this method may comprise: obtaining a sample comprising chromatin, reacting the sample with the composition to selectively label the open chromatin in the sample, and analyzing the labeled sample.

Provided herein, among other things, are various compositions and methods for analyzing chromatin. In some embodiments, the composition may comprise a mixture of a nicking enzyme, four dNTPs, at least one labeled dNTP and, optionally, a polymerase. In some embodiments, this method may comprise: obtaining a sample comprising chromatin, reacting the sample with the composition to selectively label the open chromatin in the sample, and analyzing the labeled sample.

This invention features systems and methods for the detection of analytes, and their use in the treatment and diagnosis of disease.