Regarding immunohistochemistry for colon cancer, which antigen retrieval buffer gives better results, Tris-EDTA buffer (PH 9) or Sodium Citrate buffer (PH 6)?
Depends on the antibody used. I recommend trying both of them.
Yes, I tried both of them for most cancer types such as breast cancer, liver cancer, leukaemia and different tissues such as mammary gland, liver, spleen, lung, colon, brain, skin, and kidneys. In most cases citrate buffer gave better result, except in colon cancer, in which the results with citrate buffer and Tris-EDTA buffer were the same.
you can try, sodium citrate PH3, or tris buffer saline PH 7. Also what is your antigen retrieval? You might want to play around with the amount of time you do this step.
Let me know if this works
I havn't tried PH 3 or PH 7. My antigen revivals are sodum citrate buffer, Tris EDTA buffer and Protenease K, Tried all, but still havn't got good result. Also I played with time, but still need more trying. TQVM
You can also try a steamer or pressure cooker, like Pascal (dako) or Target retrieval solution from dako. Have you tried AR without proteinase K? You can also try add tween20 0,01% in your AR solution. Are you working with a phospho protein?
Thankfully, after hard working and many trial, i got a very good result for Beta catenin expression in colon cancer. TQ for your concerning.
The success of antigen retrieval protocols in colon cancer as well as other malignant and normal tissues fixed with buffered formalise and embedded in paraffin is dependent upon the antigen (epitope) you are attempting to identify. For example both CEA and CA-19.9 are carbohydrate antigens and stable and can be more successfully exposed using low pH methods and proteases whereas high pH techniques are of little or no value with these epitopes. In contrast, protein epitopes may be successfully exposed by incubation with chelators (citrate & EDTA) in buffers pH 6 - 8. Lipid epitopes are destroyed at pH extremes but are stable with protease treatment and chelator buffers. I have attache a document that may provided options you may choose to employ should your results not be satisfactory.
Hello everyone,
I am working on pancreatic tumor, but my staining with Cytokeratin 19 doesn't work. My tissues are fixed in PFA 4%, embedded in OCT and cut by cryostat. I tried to retrieve the antigen with proteinase K, however all sections were come off ,even I used the superfrost slides and dried them overnight under the hood.
Does anyone have experience to help me in this case?
I am very thankful for your help.
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