Abstract: Background: Autofluorescence (AF)‐Raman spectroscopy is a technology that can detect tumour tissue in surgically excised skin specimens. The technique does not require tissue fixation, staining, labelling or sectioning, and provides quantitative diagnosis maps within 30 min. Objectives: To explore the clinical application of AF‐Raman microscopy to detect residual basal cell carcinoma (BCC) positive margins in ex vivo skin specimens excised during real‐time Mohs surgery. To investigate the ability to analyse skin specimens from different parts of the head‐and‐neck areas and detect nodular, infiltrative and superficial BCC. Methods: Fifty Mohs tissue layers (50 patients) were investigated: 27 split samples (two halves) and 23 full‐face samples. The AF‐Raman results were compared to frozen section histology, carried out intraoperatively by the Mohs surgeon and postoperatively by dermatopathologists. The latter was used as the standard of reference. Results: The AF‐Raman analysis was completed within the target time of 30 min and was able to detect all subtypes of BCC. For the split specimens, the AF‐Raman analysis covered 97% of the specimen surface area and detected eight out of nine BCC positive layers (similar to Mohs surgeons). For the full‐face specimens, poorer contact between tissue and cassette coverslip led to lower coverage of the specimen surface area (92%), decreasing the detection rate (four out of six positives for BCC). Conclusions: These preliminary results, in particular for the split specimens, demonstrate the feasibility of AF‐Raman microscopy for rapid assessment of Mohs layers for BCC presence. However, for full‐face specimens, further work is required to improve the contact between the tissue and the coverslip to increase sensitivity.
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