Margin-controlled staged excision (slow Mohs) has emerged as a preferred method for the treatment of lentigo maligna (LM). The interpretation of margins for LM is one of the most challenging tasks faced by a dermatopathologist. R21 is a mouse monoclonal antibody against soluble adenylyl cyclase (sAC), overexpressed in the nuclei of LM but not in native melanocytes. The objective of this study was to validate the use of sAC immunohistochemistry in histological assessment of slow Mohs surgery margins for LM. Seventeen randomly selected cases of patients who underwent slow Mohs surgery for LM at Lahey Clinic, Burlington, MA, were studied. Ninety-nine margins were stained with R21 and microphthalmia transcription factor antibodies and reevaluated blindly by 2 observers. Sixteen of 17 lesions expressed sAC. In all cases, observers agreed on interpretation of R21 stains. In 85 (86%) margins, there was concordance between routine sections and R21 stains. In 14 margins (14%), the results were discrepant. In 2 margins, R21 identified foci of LM missed on routine sections. In 8 margins, atypical melanocytes, interpreted as positive in routine sections, were negative for R21 questioning the accuracy of the original interpretation. Microphthalmia transcription factor stained nuclei of melanocytes in all margins. We found significant correlation between assessment of margins by sAC immunohistochemistry and routine histology. Evaluation of sAC expression using R21 antibody is a useful diagnostic adjunct in the evaluation of margins of LM during slow Mohs surgery.