Abstract Optical coherence tomography is a non-invasive imaging technique that enables high-resolution in vivo imaging of skin. Although optical coherence tomography is promising for diagnosing basal cell carcinoma, its limited penetration depth may impede basal cell carcinoma subtyping. This study evaluated whether topical application of glycerol can increase penetration depth and improve the image quality and visibility of characteristic features of basal cell carcinoma. A total of 61 patients with a total of 72 basal cell carcinomas were included. Optical coherence tomography scans were obtained before and after application of an 85% glycerol solution. The mean penetration depth of each optical coherence tomography scan was acquired by automatically tracing both skin surface and the point of signal loss using a custom-made MATLAB program. Mean ± standard deviation penetration depth increased from 883 ± 108 to 904 ± 88 µm before and after glycerol application, respectively (p = 0.005). Topical application of glycerol leads to a significant 2.4% increase in penetration depth. However, no significant differences in image quality and visibility of basal cell carcinoma features were found. Significance Optical coherence tomography is a promising non-invasive diagnostic technique for the diagnosis of basal cell carcinoma; however, its limited penetration depth may impede basal cell carcinoma subtyping. By reducing light scattering in optical coherence tomography scans, using optical clearing agents, such a glycerol, the penetration depth may been enhanced. This study included 61 patients with a total of 72 basal cell carcinomas. Optical coherence tomography scans were obtained before and after application of an 85% glycerol solution. Mean penetration depth increased significantly, by 2.4%, after application of glycerol.