[Antigen retrieval immunohistochemistry under the influence of pH value and time]

To find the best antigen retrieval buffer and retrieval time for the immunohistochemical staining of P504S, P63, CD10, and Ki-67.

The following tissues were fixed in 10% formalin: the tissues of prostate carcinoma, benign prostate hyperplasia, proliferative breast disease and cunnus epithelial tissue infected by human papilloma virus. Immunohistochemical stainings for these tissues were processed by EnVision's methods. The first antibodies were P504S for prostate carcinoma, P63 for benign prostate hyperplasia, CD10 for proliferative breast disease, and Ki-67 for cunnus epithelial tissue infected by human papilloma virus. The same second antibody EnVision was used for all the tissues. For each tissue, we used three different antigen retrieval buffers, CA (pH 6.0), EDTA (pH 8.0) and EDTA-Tris (pH 9.0). For each buffer, there were 4 durations for antigen retrieving, namely 12, 20, 25 and 30 min. The positive signals in different conditions were observed and compared under light microscope.

The best signal for P504S was observed after retrieval in the EDTA-Tris buffer (pH 9.0) for 20 min, P63 for 30 min, and Ki-67 for 25 min. The best positive signal for CD10 was observed after retrieval in all the buffers with different pH value for 20 or 25 min.

Different antigens have their own optimal retrieval buffers and the optimal retrieval time. Relatively, for the antigen retrieval of Ki-67, P504S and P63, EDTA-Tris buffers with pH 9.0 is better than other low pH retrieval buffers.