1. Tissue Preparation: The tissue sample is fixed and embedded in paraffin to preserve its structure. Sections are then cut using a microtome and mounted onto slides. 2. Antigen Retrieval: To unmask epitopes that may have been obscured during fixation, antigen retrieval techniques such as heat-induced epitope retrieval (HIER) are employed. 3. Blocking: Non-specific binding sites are blocked to prevent background staining. 4. Primary Antibody Incubation: The tissue sections are incubated with a primary antibody that specifically binds to the target antigen. 5. Secondary Antibody Incubation: A secondary antibody conjugated to an enzyme or fluorophore is applied, which binds to the primary antibody. 6. Detection: The signal is visualized using chromogenic substrates or fluorescence, allowing for the identification of the antigen within the tissue.
