I recently had an enquiry from one of our post docs who was looking at TUNEL staining and came across this reference in biochemica no 4 (1997)titled “Fixation of Tissue Sections for TUNEL Combined with Staining for Thymic Epithelial Cell Marker” You should be able to google it and find the article. Basically it said that after fixation with PFA to improve staining you need to treat with triton and sodium citrate in the cold to expose the antigen again. Below is the suggested protocol for thin cryostat sections. I’m sure it could be modified for thick sections.
1. Cut 4 μm thin cryosections and mount on polylysine-coated glass slides. 2. Air dry overnight at room temperature and freeze foil- wrapped slides at –20°C until use. 3. Apply frozen glass slides directly into a container with 1% buffered paraformaldehyde for 30 min at RT. 4. Rinse swiftly in PBS and immerse in a solution of 1% Triton X100 (v/v) and 1% sodium citrate (w/v) for 2 min at 4°C. 5. Wash in PBS and incubate with 50 μl TUNEL reaction mixture (TdT solution with dUTP-FITC solution, 1+9). Convert to enzyme label if desired. 6. Wash with PBS and block with dilution of adequate normal serum. 10. Proceed with desired immunohistochemical labeling. Regards Steve Weston Senior technical officer Menzies Research Institute Hobart Tasmania
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