1. Sample Preparation: Tissue sections are prepared and fixed to preserve cellular integrity. Common fixatives include paraformaldehyde or formalin. 2. Permeabilization: Samples are treated with proteinase K or other agents to permeabilize cell membranes, allowing enzyme access to DNA. 3. Labeling Reaction: TdT and labeled dUTP are added to the sample, incubating to allow the enzymatic addition of labeled nucleotides to DNA breaks. 4. Detection: Depending on the label used, detection may involve fluorescence microscopy, flow cytometry, or colorimetric methods. Fluorescent labels like FITC or Texas Red are common for microscopy, while biotin-labeled nucleotides are often used with streptavidin-HRP for chromogenic detection.
