Methods for spatial analysis using targeted rna depletion

A method for depleting an undesirable RNA molecule in a biological sample, the method comprising:

• (a) contacting the biological sample with a first substrate;
• (b) adding a plurality of undesirable RNA depletion probes to the biological sample, wherein an undesirable RNA depletion probe of the plurality of undesirable RNA depletion probes comprises a sequence that is substantially complementary to a sequence of the undesirable RNA molecule;
• (c) hybridizing the undesirable RNA depletion probe to the undesirable RNA molecule, thereby generating an undesirable RNA depletion probe-undesirable RNA molecule complex;
• (d) removing the undesirable RNA depletion probe-undesirable RNA molecule complex to deplete the undesirable RNA molecule in the biological sample;
• (e) 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 and wherein a capture probe of the plurality of capture probes comprises a capture domain;
• and
• (f) when the portion of the biological sample is aligned with at least the portion of the array, hybridizing a nucleic acid analyte or an intermediate agent from the biological sample to the capture domain of the capture probe.