TUNEL assay cell type - Mouse mammary gland tissue

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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1 year ago

1 year 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
Add Proteinase K Solution and incubate 15 to 30 minutes at
room temperature,

Add Labeling Reaction Mix and incubate at 37˚C for 1 hour in a humidity chamber.
Downstream tips
If no staining in TACS-Nuclease, increase incubation time with Proteinase K (up to 60 minutes) or Cytonin™ (up to overnight at 4 oC)
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