karl-heinz blenk (Karl at blenk.deceiver.org) wrote:
: 1) I am looking for a marker to detect apoptosis in dorsal root ganglion
: cells using light microscopy.
: I con faintly remember having heard something about Acrydyl-orange.
: Does anybody has any experience with this substance or any other
: suggestions to offer?
I'm also sending you an e-mail about this, but for the benefit of everyone
You didn't say the conditions of your experiments, but if you are planning
to fix the tissue, I'd recommend performing TUNEL or ISEL techniques for
in situ staining. TUNEL is a fairly simply procedure which relies upon
DNA fragmentation in apoptotic cells as a marker. TdT enzyme appends
base analogs to the 3OH end of DNA fragments. Next, you add a primary
antibody that tags the analogs.
It's a clever, simple technique that works pretty well (but is also fairly
ISEL is a variant of TUNEL.
You can get a TUNEL stain kit, called APOPTAG, from Oncor (USA). Ask for
the Peroxidase kit, which is a light stain. Kit cost is $300, which allows
you to do up to 50 slides (but probably 40 is more realistic). You'll also
need to develop a positive control protocol, requiring a bit of fooling around
with times of exposure. I have a protocol if you're interested.