TUNEL assay cell type - Rat pulmonary arterial smooth muscle cells

TUNEL assay is the cell death detection method where the biochemical marker of apoptosis is DNA fragmentation. The assay involves the microscopical detection of generated DNA fragments with free 3'-hydroxyl residues. in apoptotic cells using enzyme terminal deoxynucleotidyl transferase (TdT) which adds biotinylated nucleotides at the site of DNA breaks. Major challenges of this method involve proper access of the enzyme which could be hampered by poor permeabilization and/or excessive fixation with cross-linking fixative (common with archival tissue). This issue can be resolved by optimizing the incubation time with Proteinase K or CytoninTM.

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2 years ago

2 years ago by Ercole Udinesi Italy

Are 10 week old samples in formalin still usable?

I have left my tumor samples in formalin for an extended period of time (around 10 weeks). Do you think I will be able to use them for TUNEL assay and get results? Thank you for your help in advance

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Protocol tips
- Slides were fixed with 4% paraformaldehyde for 30 min at room temperature.

- Slides were permeabilized with 1% Triton X-100 for 4 min at 4°C instead of 2min on ice.

- Slides were labelled at 37°C for 60 min in a dark environment
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