Objective To investigate the clinical significance of gene methylation of p53-bax mitochondrial apoptosis pathway in the carcinogenesis of cholangiocareinoma. Methods Promoter hypermethylation of DAPK, P14^ARF and ASC genes were detected by methylation-specific PCR. Exon 5-8 of p53 gene were examined by automatic sequencing. Results It was found that 66.7% of 36 cholangiocarcinoma patients had methylation of at least one tumor suppressor gene. The rate of tumor suppressor gene methylation in these cholangiocareinomas was 25.0% in P14^ARF, 30.6% in DAPK and 36. 1% in TMS1/ASC. The methylation rate of tumor suppressor gene in tissues adjacent to the cancer tissue was 13.9% including 5.6% in DAPK and 8.3% in TMS1/ASC. p53 gene mutation was detected in 22 of 36 patients（61.1% ）. Fourteen patients （38.9%）was found to have p53 gene mutation associated with the methylation of tumor suppressor gene. The rate of p53 gene mutation and methylation of tumor suppressor gene were statistically and significantly correlated with the features of tumor biology including differentiation and invasion （P 〈 0.05）. Conclusion DNA methylation of p53-bax mitchondfial apoptosis pathway may be a frequent molecular event in the carcinogenesis of cholangiocarcinoma. Although the methylation rate of ASC, DAPK genes is relatively low, it may be helpful for early diagnosis, p53 gene mutation associated with the methylation of tumor suppressor genes may be correlated with tumor malignant biologic features.