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Grading melanocytic dysplasia in paraffin wax embedded tissue by the nucleic acid index

¹ Laboratory of Pathology, National Cancer Institute, Bldg 10-2N212, 10 Center Drive Bethesda, MD 20896, USA
² Laboratory of Experimental Carcinogenesis, National Cancer Institute, Bethesda, MD 20892, USA

Correspondence to:
Dr D M Berman
Bristol-Myers Squibb, Princeton, NJ, USA; david.berman{at}bms.com Background: Although nucleic acid derangements are the hallmark of melanocytic dysplasia, the gold standard for its diagnosis remains the microscopic evaluation of haematoxylin and eosin stained slides. However, light microscopy is subjective and crucial genomic changes do not always show as changes in histology.

Aims: To introduce the nucleic acid index (NAI) as a means of analysing nucleic acid derangements in histological sections at the level of the individual cell and within the context of its microenvironment.

Methods: Confocal laser scanning microscopy was performed on melanocytic lesions stained with acridine orange (AO), a fluorescent stain for DNA and RNA. The NAI, calculated by measuring the fluorescence intensities of AO in nuclei relative to the surrounding cytoplasm, reflects the concentration of DNA relative to RNA.

Results: When applied to benign naevi, dysplastic naevi, and melanoma, a very strong significant association was seen between lower NAI and malignant potential (p < 0.0001). Strong inverse correlations were found between NAI and both mitotic index and Breslow thickness. Interestingly, the NAI for dysplastic naevi is between that of melanoma and most benign naevi, consistent with their intermediate biological behaviour and histological appearance.

Conclusion: By providing a quantitative measure for melanocytic neoplasia, the NAI may improve the diagnosis of melanocytic lesions and the selection of treatment.

Abbreviations: AO, acridine orange; CLSM, confocal laser scanning microscopy; DAPI, 4',6-diamidino-2-phenylindole; DMN, dysplastic melanocytic naevi; FFPE, formalin fixed, paraffin wax embedded; H&E, haematoxylin and eosin; NAI, nucleic acid index; PBS, phosphate buffered saline; SR, surface rendering

Keywords: melanoma; dysplasia; confocal; nucleic acids; acridine orange