Cleavable fluorescent tyramide for sensitive and multiplexed analysis of biological samples

A method of multiplexed in situ analysis of biomolecules in a tissue, the method comprising the following steps:

• (a) performing a first contacting step that comprises contacting a tissue comprising a plurality of biomolecules to cleavable detectably-labeled tyramide, wherein the first contacting step occurs under conditions that promote conjugation of the cleavable fluorophore-labeled tyramide to a target biomolecule;
• (b) performing a second contacting step that comprises contacting the tissue with a plurality of horseradish peroxidase (HRP)-conjugated targeting agents that are configured to specifically bind or hybridize to the target biomolecule in the contacted tissue, wherein the second contacting step occurs under conditions that promote binding or hybridization of the targeting agents to the target biomolecule;
• (c) imaging the cell after the second contacting step whereby a detectable signal generated from an interaction of HRP-conjugated targeting agents with the cleavable detectably-labeled tyramide is detected;
• (d) removing the fluorophore from the fluorophore-labeled tyramide;
• and
• (e) optionally consecutively repeating the contacting, imaging, and removing steps, each time with a new plurality of HRP-conjugate targeting agents for each subsequent cycle, wherein each utilized plurality differs from each other utilized plurality due to being configured to specifically bind or hybridize to a different target biomolecule.