The normal oral mucosa from 77 individuals of known smoking and alcohol-intake history was cultured. After 14-21 days in culture, epithelial cells were stained for p53 expression using immunohistochemistry. PCR-SSCP analysis of the p53 gene was also performed on a random sub-set of these samples (20 non-smokers and 21 smokers). Expression of the stable, non-functional form of p53 protein as detected by the p53-240 antibody was found to be significantly elevated in the cultured oral mucosa of smokers. (P < 0.01). PCR-SSCP analyses indicated a higher level of base changes in smokers than in non-smokers. These observations are consistent with other findings of significantly increased p53 protein expression in the oral mucosa and other tissues of smokers and suggests that p53 mutations may be an early event in smoking-induced oral cancers.