Abstract: Objective : To develop an ideal method for detection of the integration status of HPV16 for clinical use and to explore the role of HPV16 integration in cervical carcinogenesis and its potential clinical significance. Methods : Multiplex real-time PCR was employed to quantify the copy number of E2 and E6 genes and analyze the integration status of HPV16 DNA according to E2/E6. The integration status of 49 cases of cervical intraepithelial neoplasia (CIN) and 51 cases of squamous cell cervical cancer that tested positive for HPV16 was investigated using paraffin-embedded tissues. Results : a) The correlation coefficients for E2 and E6 standard curves were 1.0 and the amplification efficiencies were both above 95%. b) The cutoff value of E2/E6, used to distinguish the pure episomal form from a mixed form of episomal and integrated HPV16 DNA, was 0.81 in the multiplex real-time PCR test. c) HPV16 existed as episomes in most of the CIN Ⅰ lesions, and 42.9% of them had mixed episomal/integrated HPV16. The integrated form of HPV16 constituted the majority in most of the CIN Ⅱ and CIN Ⅲ cases. In squamous cervical carcinomas, the HPV16 was mostly integrated into the host chromosome. d) The rate of HPV16 integration was associated with the degree of cervical lesions and the rate increased with the progression of cervical disease. e) The rate of pure integrated HPV16 in stage Ⅱand Ⅲ (88%) was higher than that in stage Ⅰ (33.3%). Conclusions : a) Multiplex real-time PCR is a rapid and sensitive method for detection of the integration state of HPV16 DNA. b) The integration of HPV16 DNA is a very early event in the development of cervical cancer and may act as an activation mechanism for progression from preinvasive to invasive cervical cancer. c) The pure integrated HPV16 in cervical cancer may be a useful prognostic indicator.