A microwave method that enhances detection of aberrant p53 expression in formalin-fixed, paraffin-embedded tissues

Abstract

Objective: Detection of aberrantly accumulated p53 protein by immunohistochemistry may have prognostic significance in many human neoplasms. We wished to identify a technique applicable to formalin-fixed, paraffin-embedded sections that would reliably yield results equivalent to frozen-section immunohistochemistry.

Design: We compared the frequency of p53 immunostaining obtained by applying monoclonal antibodies PAb1801, DO7, or DO1, a 1:1 PAb1801-DO7 cocktail, and a 1:1 PAb1801-DO1 cocktail to fresh-frozen and formalin-fixed, paraffin-embedded tissues from 36 lung and upper aerodigestive-tract carcinomas. With the formalin-fixed tissues, we compared pepsin predigestion with microwave irradiation in citrate buffer as means of enhancing the sensitivity of p53 detection.

Setting and patients: All tissues were obtained from surgical resections of tumors, from patients who underwent surgery at the Minneapolis Department of Veterans Affairs Medical Center between 1990 and 1992.

Main outcome measures: The sensitivity of different paraffin section techniques for immunohistochemical detection of accumulated p53 protein was determined in reference to the optimal frozen section method (defined as the method that yielded the greatest number of p53-positive cases in frozen tissue).

Results: Microwave antigen retrieval markedly enhanced staining with PAb1801 and DO7 in paraffin sections, as compared with pepsin predigestion and no pretreatment. This technique was 100% sensitive relative to the optimal frozen tissue method. In contrast, staining with DO1 alone was not enhanced by microwaving.

Conclusions: Microwave pretreatment in conjunction with the use of either PAb1801 or DO7 is highly efficacious in the immunohistochemical detection of aberrant p53 expression in formalin-fixed, paraffin-embedded tissues.