The process of IHC staining involves multiple steps:
Tissue Preparation: The tissue is fixed, usually with formalin, and then embedded in paraffin to preserve its structure. Sectioning: Thin sections of the tissue are cut and placed on slides. Antigen Retrieval: To unmask antigens, the tissue sections are treated with heat or enzymes. Blocking: Non-specific binding sites are blocked to prevent background staining. Primary Antibody Incubation: The tissue is incubated with a primary antibody specific to the target antigen. Secondary Antibody Incubation: A secondary antibody that binds to the primary antibody is applied. This secondary antibody is usually conjugated to an enzyme or fluorophore. Detection: The enzyme or fluorophore reacts with a substrate to produce a colorimetric or fluorescent signal, indicating the presence of the antigen.
