Reply

We appreciate the comments of Tuccari et al regarding our recent study of argyrophilic staining of nucleolar organizer region (AgNOR) counts and morphometry in benign and malignant melanocytic lesions. In this study, we found that both AgNOR counts and AgNOR morphology help to distinguish benign from malignant melanocytic lesions and that the combination of AgNOR counts and morphometric parameters can improve the discriminating ability of the technique. 1 We agree that the AgNOR staining and determination under strictly controlled and preferably uniform conditions will assist in obtaining results that will be comparable among different investigators. We also agree that the use of an image analysis system should help to facilitate this. As pointed out by Tuccari et al and by many others, AgNOR quantification can readily be affected by many factors, including the method of tissue fixation, staining conditions such as temperature and duration, whether an inverted incubation technique is employed, and the methodology applied to determine AgNOR measurements. The bleaching procedure employed in the study of melanocytic lesions is also an important factor that may affect the AgNOR quantification. 2 The application of a standardized AgNOR method, such as that recommended by the Committee on AgNOR Quantitation within the European Society of Pathology, 3 may assist in producing results that are more comparable among different laboratories. However, the presence of melanin pigment within melanocytic lesions will probably limit the usefulness of the methodology recommended by Tuccari et al in assessing AgNOR variables in melanocytic lesions, without the use of a prior melanin bleach procedure such as the one we described. 2 As we have previously demonstrated, 1,2 our procedure has been very successful for assessing both AgNOR counts and AgNOR morphology in melanocytic tumors. It may be that specific standardized AgNOR methods are required for the evaluation of certain tumors. It would thus be valuable if a melanin bleach procedure is incorporated into the standardized AgNOR methods recommended by the Committee on AgNOR Quantitation within the European Society of Pathology 3 for the assessment of melanocytic lesions. Although we agree with Tuccari et al that the AgNOR method has been shown to have prognostic significance in a variety of tumors, 4,5 the purpose of our work was to evaluate the diagnostic utility of AgNOR method in distinguishing benign nevi from malignant melanomas. 1 While we acknowledge that the diagnostic value of individual AgNOR markers may be limited by the phenomenon of overlapping values between benign and malignant lesions, the use of a combination of AgNOR counts and morphometric parameters may improve the diagnostic value of this technique in melanocytic lesions. 1 Our multivariate studies have suggested that the AgNOR diagnostic values in melanocytic lesions may be improved further when co-evaluated with other cytometric markers, such as DNA microdensitometry and MIB1-Ki67 immunoreactivity. 6 Further evaluations of these techniques are required in difficult melanocytic lesions that cannot readily be diagnosed as benign or malignant using standard histopathologic methods. The prognostic significance of the AgNOR technique in melanoma patients using our protocols is yet to be investigated and also requires further study. Ling-Xi L. Li, BM, PhD Richard A. Scolyer, MBBS, FRCPA Kerry A. Crotty, MBBS, BSc(Med), FRCPA Allan A Palmer, MBBS, FRCPA Jillian J. Kril, BSc(Hons), PhD John F. Thompson, MD, FRACS, FACS Stanley W. McCarthy, MBBS, FRCPA