I found this paper using Citraconic Anhydride and Heat (please see attached) and I would like to know if anyone has successfully replicated it.
Hi Madhur, you may check on this link. In my case, I used to heat my paraffin section in heated citrate buffer, pH 6 AT 100C for 20 mins and managed to observe the expression. However it depends to your AB as some AB no need any AR. You need to optimize it. Good luck!
https://www.researchgate.net/post/Antigen_retrieval_for_immunofluorescent_staining/2
I had good results using pressure cooker (12 minutes) and water bath (30-40 minutes) techniques in the past. I used citrate pH 6 and EDTA pH 9 buffers. Another method that can be used is by boiling the slides in a microwave for 6 minutes two times.
Personally, I think water bath is the best Antigen Retrieval technique I have tried.
Dear Khairunnisa, Mohamed and Elena,
Thank you very much for taking time to answer my question.
We had success with protocols based on your suggestions and as rightly pointed out, diferent Abs require different retrieval techniques.
However, I am interested to stain a FFPE section/slide with two/three different Abs (and then perform IF) to understand co-localizatoin/co-expression and unfortunately, each of my Ab of interest requires different antigen retrieval technique. To circumvent this, I was wondering if there is any 'single' technique (in line with the paper I have attached, which uses citraconic anhydride and heat) that can work for all the antibodies and have users here tried it with good luck.
Best wishes,
Maddy.
Dear Madhur, that was interesting. I am looking for that too as my 2 ABs required heat ARA and without heat AR. Looks so challenging. Unable to co-stain yet. Thanks for the question.
https://www.ncbi.nlm.nih.gov/pubmed/?term=luchini+cd71
https://www.ncbi.nlm.nih.gov/pubmed/?term=luchini+twist1
- Badges
- Science method