Optical coherence tomography (OCT) can be used as an adjunct to colposcopy in order to detect precancerous and cancerous cervical lesions. Optical clearing agents (OCAs) temporarily reduce the optical scattering of biological tissues. The purpose of this study was to investigate their influence on OCT imaging. OCT images were taken from unsuspicious and suspicious areas of fresh conization specimens immediately after resection and 5, 10, and 20 min after application of dimethyl sulfoxide (DMSO) or polyethylene glycol (PEG). Corresponding histologies were obtained from all sites. The images taken 5, 10, and 20 min after application of OCA were compared to the initial images with respect to changes in brightness, contrast, and scanning depth using a standard nonparametric test of differences of proportions. Further, mean intensity backscattering curves were calculated from all OCT images in the histological groups CIN2, CIN3, inflammation, and normal epithelium. Mean difference profiles within each of these groups were determined, reflecting the mean differences between the condition before the application of OCA and the exposure times 5, 10, and 20 min, respectively. The null hypothesis was tested employing the Dicky-Fuller-test, Hotelings-test and run test. The visual analysis of 434 OCT images from 109 different sites of 24 conization specimens showed a statistically significant increase in brightness and contrast for normal and dysplastic epithelium after application of DMSO or PEG. Further, the analysis of mean intensity profiles suggests the existence of an increased backscattering intensity after application of DMSO or PEG. DMSO and PEG contribute substantially to optical clearing in cervical squamous epithelium and therefore influence OCT imaging in a positive way. With further refinement of the OCT technology, the observed changes may be beneficial in interpreting the tissue microstructure and identifying cervical intraepithelial neoplasia.