Hello everyone, hope you are doing well.
I have FFPE brain arteriovenous malformation (AVM) tissues I have been trying to stain with cleaved-caspase-3.
I have used the Asp175 (D3E9) CC3 9579 from cell signaling at: 1:100 and 1:50. However, I am not able to detect any signal from that channel while my other channels are expressing the co-stains I used.
Troubleshootings I have tried:
-I have been diluting Ab in 10%DS (which has worked effectively for my other abs)
-Antigen retrieval at low pH
-antigen retrieval at high pH
-antigen retrieval using pressure cooker as well as overnight.
-Longer primary incubation time
Any recommendations would be greatly appreciated!
best
Hello,
We have in the past used Cleaved Caspase-3 (Asp175) Antibody #9661 from Cell Signaling Technology, and worked well on FFPE tissue after heat-induced antigen retrieval in low pH buffer
I would recommend combining antigen retrieval methods, high pH and temperature, in addition to concentrating the primary antibody.
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