Abstract: Objective: To investigate the role of p16/Ki-67 double-staining in diagnosis and grading of cervical intraepithelial lesions. Methods: Immunohistochemical dual stain was used to detect the co-expression of p16 and Ki-67 in 15 cases of cervical immature metaplasia (IM), 32 cases of low-grade squamous intraepithelial lesion（LSIL, CIN Ⅰ）, 90 cases of high-grade squamous intraepithelial lesion (HSIL, CIN Ⅱ and CIN Ⅲ). The correlation of p16 and Ki-67 co-expression with diagnosis and grading of cervical intraepithelial lesion was analyzed. Results: p16 was negative in IM, and was of focal or patchy staining with weak intensity and positive cells restricted to the lower third of squamous epithelium in CIN Ⅰ. In HSIL, diffuse p16 positive immunostaining cells of 2/3 or full thickness of squamous epithelium was seen. The positive rates of p16 protein were 6.67%, 62.50%,88.89%, 95.56% in IM, CIN Ⅰ, CIN Ⅱ,CIN Ⅲ, respectively, and the differences between these 4 groups were statistically significant (P<0.05). For Ki-67, expression only existed in the basal or parabasal cells in IM. In CIN Ⅰ, Ki-67 immunostaining was restricted to the lower third of the squamous epithelium, while diffuse Ki-67 immunostaining in 2/3 or full thickness of the squamous epithelium was seen in HSIL. The Ki-67 indices were 7.21%, 16.41%, 33.54%, 50.32% in IM,CIN Ⅰ, CIN Ⅱ, CIN Ⅲ, respectively, and the differences were statistically significant (P<0.05). Dual staining did not lead to the absence of Ki-67-positive signal. Ki-67 index was 36.47% in p16 positive region, while was 10.53% in p16 negative region. With the increase in grading of cervical epithelial lesion, the cases of p16/Ki-67 co-expression increased. The co-expression rates of p16/Ki-67 were 0, 51.35%, 85.45%, 99.42% in IM、CIN Ⅰ、CIN Ⅱ、CIN Ⅲ, respectively (P<0.05). Conclusions: The expression of p16 and Ki-67 is associated with the increase in grading of cervical squamous intraepithelial lesion. There is a significantly positive correlation between the expression of p16 and Ki-67. Immunohistochemical dual staining is helpful for the diagnosis and grading of cervical intraepithelial lesions.

Key words: Cervical squamous intraepithelial lesion, Immature metaplasia, Immunohistochemistry dual stain, Protein,p16, Protein,Ki-67