Immunohistochemical demonstration of p53 protein in colorectal adenomas and adenocarcinomas. Reliable application of the heat‐induced antigen retrieval method to formalin‐fixed, paraffin‐embedded material

Ninety‐nine polypoid neoplasms and eight advanced adenocarcinomas of the colon were studied immunohistochemically for p53 protein expression. For reproducible antigen retrieval, formalin‐fixed, paraffin‐embedded archival sections were heated at 90°C for 120min in 0.01 mol/L phosphate‐buffered saline, pH 7.2, prior to immunostaining. Proliferating cell nuclear antigen served as a positive control marker for effective antigen retrieval. The 99 polyps were categorized into 24 high‐grade adenomas, 60 non‐invasive cancer‐in‐adenomas (CIA), and 15 CIA with stromal invasion. All the polyps contained portions of low‐grade adenoma. Positive nuclear staining of p53 protein was observed in none of the non‐neoplastic mucosa, nine (9%) of 99 low‐grade adenomas, 17 (71%) of 24 high‐grade adenomas, 46 (77%) of 60 non‐invasive CIA, 10 (67%) of 15 invasive CIA, and five (63%) of eight advanced carcinomas. When the antigen retrieval treatment was omitted, the positivity rates were 0, 2, 17, 35, 40, and 63%, respectively. When the antigen‐retrieved staining pattern was classified into (i) ‘sparse’ (< 25% of the nuclei of neoplastic glands labeled), ‘scattered’ (25–75%) and ‘dense’ (> 75%); or (ii) ‘focal’ (the positively labeled glands occupying < 25% of the tumor area), ‘intermediate’ (25–75%) and ‘diffuse’ (> 75%), the sparse and focal patterns predominated in high‐grade adenomas and non‐invasive CIA with low‐grade atypia, while the dense and diffuse patterns predominated in invasive CIA and all the advanced carcinomas revealed the dense and diffuse patterns. Non‐invasive CIA with high‐grade atypia belonged to an intermediate type between the two groups. These findings indicate that accelerated intranuclear accumulation of immunoreactive p53 protein is closely correlated with colorectal tumorigenesis.