IHC

Immunohistochemistry (IHC) is a laboratory technique used to detect specific antigens in tissue sections by leveraging the binding of antibodies to their targets. It provides anatomical context for molecular findings and is widely used for diagnosis, classification, prognosis, and treatment planning in pathology and research. IHC is commonly performed on formalin-fixed, paraffin-embedded tissues, though frozen sections may be used for certain antigens.

In a typical IHC workflow, tissue sections are prepared and may undergo antigen retrieval to unmask epitopes. Blocking steps reduce non-specific binding, after which a labeled antibody is applied. In direct methods, a labeled primary antibody binds the antigen; in indirect methods, a secondary antibody against the primary antibody carries a detection label. Detection reagents linked to enzymes such as horseradish peroxidase or alkaline phosphatase generate a chromogenic signal visible under light microscopy, often producing a brown or red precipitate. Fluorescent labels enable multiplexing and visualization by fluorescence microscopy. Proper controls, including positive and negative samples, are essential to validate results.

IHC is used to identify cell lineage and tumor type, determine protein expression patterns, and inform therapeutic decisions. Common clinically important targets include hormone receptors (such as estrogen and progesterone receptors), HER2/neu, proliferation markers (Ki-67), and immune checkpoint proteins (e.g., PD-L1). The technique also serves in basic and translational research to localize proteins within tissue architecture.

Key considerations include antibody specificity and validation, pre-analytic variables (fixation and processing), standardized staining protocols, and careful interpretation of staining localization (nuclear, cytoplasmic, or membranous), intensity, and extent. Limitations involve cross-reactivity, false positives or negatives, and challenges in standardization across laboratories.