The aim of this study was to investigate the heat stability of squamous cell carcinoma (SCC) antigen, a tumor-associated serine proteinase inhibitor (serpin) in tumor tissue, extract by electrophoretic methods. After heat treatment at 70 degreesC for 2 h, the tumor tissue extract showed a single main protein band of 45 kDa on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) which reacted with a monoclonal antibody specific for SCC antigen. The heat-stable SCC antigen was separated by two-dimensional electrophoresis (2-DE) into four spots with p/ 6.4-5.9 and M-r 44500-45000 of SCC antigen-1. Furthermore, the SCC antigen-1 still showed its inhibitory activity against a cysteine proteinase, papain, by gelatin zymography. These results suggest that heat treatment of protein sample at 70 degreesC for 2 h may be a useful method for a partial purification of SCC antigen-1 which can inhibit lysosomal cysteine proteinases such as cathepsin L S and K.