Apoptosis, Necrosis or Autophagy?

[admin]

Hi again Jpap,

really excellent post on the issue!

I also agree, late stages are difficult to separate out however newer methods are making this possible.

[jpap]

Thank you for your kind words Admin, I am happy to share the few things I know on the issue.

[Yak-Nam Wang]

Hi,

I have a question following this thread. Ideally I would like to determine the mecanism of cell death in treated porcine pancreatic tissue. However, due to the difficulties in this, I opted for the 'easier' option of determining if death was via apoptosis.

I have porcine pancreatic tissue that was treated (non-chemical) in vivo. Biopsies were taken 7 days after treatment.

Looking at sections stained with H&E I can see pyknotic nuclei and nuclear fragments which would suggest apoptosis. However, when labelling for activated caspase-3 (cell signalling) I don't see anything labelling. I chose caspase-3 first before doing an end-nick assay due to the controversy behind the latter.

As a side note, pigs do not have tonsils so it is possible that my positive controls are off (I used pig placenta). Although, my negative controls (DAN only, No ab, no primary) come out 'clean' and my placenta has distinct labelling.

Regarding your mention of the correct time line, I was thinking that I have missed the stage when I can detect activated caspase-3 and instead am in secondary necrosis. Does anyone have and suggestions regarding identifying late stage apotosis or any other suggestions?

Any insight would be greatly appreciated!
Thank you.