What are the Steps Involved in Immunohistochemistry?
The immunohistochemistry process involves several steps:
1. Tissue Preparation: Tissue samples are fixed, usually with formalin, to preserve their structure and proteins. They are then embedded in paraffin, sectioned, and mounted on slides.
2. Antigen Retrieval: Fixation can mask protein epitopes, so antigen retrieval is performed to unmask them. This can be achieved using heat or enzymatic treatment.
3. Blocking: Non-specific binding sites are blocked to prevent background staining. This is typically done using a solution containing proteins like bovine serum albumin (BSA) or serum from the same species as the secondary antibody.
4. Primary Antibody Incubation: The tissue sections are incubated with a primary antibody specific to the target protein.
5. Secondary Antibody Incubation: A secondary antibody, conjugated with a detection marker, binds to the primary antibody.
6. Visualization: The detection marker, often an enzyme or fluorophore, produces a colorimetric or fluorescent signal that can be visualized under a microscope.
