COMPARISON OF P53 IMMUNOREACTIVITY IN FRESH-CUT VERSUS STORED SLIDES WITH AND WITHOUT MICROWAVE-HEATING

Because of technical problems in immunohistochemical staining of archi val material, antigens can be masked or lost. A recent study reported diminished p53 immunoreactivity in slides that had been sectioned from parafinn-embedded tissue blocks and stored at room temperature. To in vestigate this issue, we performed immunohistochemical staining with u se of a p53 monoclonal antibody (DO7) in 13 head and neck squamous cel l carcinoma (HNSCC) and 13 non-small-cell lung carcinomas. The fresh-c ut and stored slides were simultaneously stained with and without the use of a microwave heating (MWH) technique, and we compared the result s of p53 immunostaining. The stored slides were sectioned from paraffi n blocks 4 to 25 years old and kept at room temperature for 6 to 48 mo nths. The slides were blindly evaluated for percentage of positivity a nd staining intensity. Twelve HNSCCs and six lung carcinomas showed p5 3 positivity. The stored slides showed a considerable decrease in stai ning intensity (P = 0.039), compared with fresh-cut slides. The differ ence in the percentage of positivity between the stored and fresh-cut slides was statistically significant, but the mean value of the differ ence was only 3.6%, which might not be meaningful for semiquantitation of immunostaining. MWH greatly enhanced staining intensity and percen tage of positivity for both stored and fresh-cut slides. When MWH was applied, no significant difference in staining intensity (P = 0.063) w as detected in fresh-cut versus stored slides, but the difference in t he percentage of positivity was statistically significant (mean value, 3.1%). Individual cases showed a consistent p53 status regardless of the MWH treatment, storage duration, or age of the blocks, This study demonstrated a considerable decrease in p53 immunoreactivity in stored slides. Because the MWH successfully retrieved the p53 antigen withou t causing a change in p53 status, stored slides combined with an MWH a ntigen retrieval technique in a metal-containing solution should provi de p53 immunostaining results similar to those from fresh-cut slides, as long as staining intensity is not a sole study parameter.