[Histonet] TUNEL assay validity issues

[Seema Shroff]

Hi everyone,

 

I am a grad student doing fluorescently labeled TUNEL assay on mouse spinal
cord frozen sections. I used 10 day mouse liver as my positive control and
I'm pretty sure that my technique works. But I have a question - The TUNEL
enzyme does not distinguish between apoptotic DNA breaks and DNA physically
broken (especially on the surfaces of the section) due to the cryostat knife
slicing through the tissue block.

 

The Roche technical support confirmed this. TUNEL is such a widely used
technique and well-accepted. Can anyone shed light on this doubt? Is it
really of concern and if yes, what can I do to validate this method?

 

Thank you,

 

Seema.