Immunohistochemical antigen retrieval precautions

Some tissue cells fixed by formaldehyde may block some antigenic determinants due to the formation of aldehyde bonds or carboxymethyl groups during fixation, which may significantly reduce the sensitivity of immunohistochemical labeling. Therefore, some antigens need to be repaired or exposed before staining. .

Antigen retrieval methods can be divided into chemical methods and physical methods. The chemical method is enzymatic digestion method, commonly used trypsin and pepsin, and the concentration and digestion time should be moderate. Common physical methods are simple heating, microwave processing and high pressure heating. When these three methods are used, the ionic strength and pH of the soaked section buffer, the temperature and time of heating all affect the antigen retrieval effect.

The pH of the antigen retrieval solution is very important, and the effective antigen retrieval pH is more important than the chemical composition of the repair solution. The same repair solution gradually increases in intensity with increasing pH, but the optimal pH range is 6.0-10.0. The best antigen-repairing solution currently recognized is citrate buffer pH 6.0 and EDTA buffer pH 8.0. As a general-purpose repair solution, the alkaline pH of the repair solution is more effective than the acid, and for the old archived tissue that has been fixed for a long time, the acidic pH repair solution is superior to the alkaline repair solution.