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Previously, increased expression of tight junction protein claudin-1 (CLDN1) was detected in premalignant and malignant cervical lesions and applied for cytology screening. To improve the specificity, a double immunoreaction with CLDN1/Ki67 was developed in the recent study. Parallel p16/Ki67 (CINtec(R) PLUS) and CLDN1/Ki67 dual-stained cytology and histology were performed and compared. p16/Ki67 immunoreaction showed positivity in 317 out of 1596 smears with negativity in 1072 and unacceptable reactions in 207 samples. CLDN1/Ki67 dual staining was positive in 200 of 1358 samples, negative in 962, whereas 196 smears could not be evaluated due to technical reasons. Considering the high-grade squamous intraepithelial lesion cytology as gold standard, sensitivity of CLDN1/Ki67 reaction was 76%, specificity was 85.67%, while for p16/Ki67 sensitivity was 74% and specificity was 81.38%. Comparison of CLDN1/Ki67 and p16/Ki67 dual stainings showed the results of the two tests not to be significantly different. Analysing histological slides from 63 cases, the results of the two tests agreed perfectly. 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