After some paper-reading, I realized there are two ways to quantify TUNEL: either by flow cytometry, or using chamber slides or cytospin to adhere cells to a glass slide, performing TUNEL with an in-situ kit, and then counting by direct microscopy.
When I presented this discrepency to a senior lab member, he just gave me a cold stare and asked me which was more accurate: counting 400 cells or 10,000? However, I think there must be some reason why so many papers use the glass slide method !
Does it have something to do with the adhesiveness of cells (all the papers using glass slides use adhesive cells)? After all that failure and disappointment I went through with annexinV/PI, I don’t want to make the same mistake again!
And another question: If I’m doing TUNEL via flow, is single staining enough or should I counter-stain it with PI or something? I’m not sure what the function of counter-staining in this situation actually is…. Anyways I’m a newbie in this apoptosis thing (my prof asked me to do this two months ago), any help at all would be greatly appreciated!