Hello, I will give you this technique that we use, however there is one with antibodies and another without antibodies, I will provide you with the one that uses monoclonals, which is the most sensitive, but if you are interested, I can give you another one that, as I indicated, does not use monoclonals, no. It is so specific but it is very beautiful to do and see.

Requirements and Materials:

• Paraffin-embedded tissue sections, 4-5 microns thick. (BE CAREFUL, MAKE SURE IT'S PARAFFIN AND YOU DON'T PARAGRAPH OR COMPLICATE THINGS.)
• Primary antibody: Anti-tryptase (commercially available).
• Secondary antibody conjugated with enzymes (e.g., horseradish peroxidase or alkaline phosphatase) or fluorochromes.
• Buffer solutions for washing (PBS, TBS, etc.).
• Protein blocker (e.g., normal serum or 1% bovine serum albumin in PBS).
• Antigen retrieval solution (e.g., 10 mM citrate buffer, pH 6.0).
• 3% hydrogen peroxide solution (if using horseradish peroxidase as the secondary enzyme).
• Chromogenic substrate solution or staining solution for detection.
• Hematoxylin (for nuclear contrast).
• Xylene or equivalents for slide mounting.

Procedure:

This is a basic IHC protocol for detecting tryptase in mast cells. However, it's important to consult the specific recommendations of the antibody manufacturer and adapt the protocol to your laboratory conditions and available equipment. Additionally, adhere to proper safety measures when working with chemicals and biological reagents.

Thank you, Alberto. I will try it ASAP. I was more curious about details and caveats though. Like the type of unmasking buffer or if I should use a rabbit serum