Melanocytic lesions: A pilot study using dual HMB-45/Ki67 staining with a new depth measurement to aid diagnosis


Lancaster depth
Breslow depth


Diagnostic pitfalls have been widely documented on atypical naevi, a group of pigmented lesions that occupy a grey portion between benign and malignant variants. As malignant melanoma is associated with a high mortality rate it is vital we have a reliable diagnostic testing method for these challenging lesions. A pilot study was proposed using dual immunohistochemistry staining with a new depth measurement, the Lancaster depth, to develop a validating tool to measure difficult atypical lesions against. The Lancaster depth was recorded for 68 pigmented lesions; 34 benign and 34 malignant, where measurement was taken from the granular layer to the deepest HMB-45/Ki67 dual stained melanocyte. This measurement was compared against the established Breslow depth measurement where the distance between the two was of interest. The mean malignant Lancaster depth at 1.51mm was higher than that of the benign at 0.12mm. There was a statistically significant difference of 1.40mm, P = <.001. There was a statistically significant difference between the mean Lancaster and Breslow depths in both malignant and benign lesions. In the malignant group this was 0.28mm, P = <.001 compared to the benign which was 1.07mm, P = <.001. A narrow distance ratio between the two depths indicated a malignant diagnosis. A wider distance ratio was indicative of a benign diagnosis. As a diagnostic validator the Lancaster depth shows promises but has limitations as a standalone model. Working in conjunction with the Breslow depth we potentially have a useful adjunct to accurately diagnose difficult atypical lesions.


Cancer Research UK (2017). Skin Cancer Mortality.

Cichorek M, Wachulska M, Stasiewicz A, Tymi A. Skin melanocytes: biology and development. Advances in Dermatology Allergology 2013;30(1):30-41. Available from:

Bastian BC. The molecular pathology of melanoma: an integrated taxonomy of melanocytic neoplasia. Annual Review of Pathology 2014;9:239-271. Available from:

Gundalli S, Kadadavar S, Singhania S, Kolekar R. Histopathological spectrum of benign melanocytic nevi – our experience in a tertiary care centre. Our Dermatology Online 2016;7(1):21-25.

Haenssle HA, Mograby N, Ngassa A, et al. Association of patient risk factors and frequency of nevus-associated cutaneous melanomas. JAMA Dermatology 2015; Nov 4:1-8. Available from:

Batus M, Waheed S, Ruby C, et al. Optimal management of metastatic melanoma: current strategies and future directions. American Journal of Clinical Dermatology 2013;14(3):179-194. Available from:

Uguen A, Talagas M, Costa S. et al. A p16-Ki-67-HMB45 immunohistochemistry scoring system as an ancillary diagnostic tool in the diagnosis of melanoma. Diagnostic Pathology 2015;10:195. Available from:

Karimkhani C, Green AC, Nijsten T. et al. The global burden of melanoma: results from the Global Burden of Disease Study 2015. British Journal of Dermatology 2017;177(1):134-140. Available from:

Brehmer F, Ulrich M, Haenssle HA. Strategies for early recognition of cutaneous melanoma – present and future. Dermatology Practical & Conceptual 2012;2(3). Available from:

Gilmore S. Melanoma screening: informing public health policy with quantitative modelling. PLoS ONE 2017;12(9). Available from:

Patterson JW, Hosler GA. Lentigines, nevi and melanomas. In: Patterson JW. Weedon’s skin pathology, 4th ed. Oxford: Elsevier, 2016:888.

Calonje E, Brenn T, Lazar A, McKee PH. McKee’s pathology of the skin: with clinical correlations. 4th ed. Oxford: Elsevier/Saunders, 2012.

Rebecca VW, Sondak VK, Smalley KSM. A brief history of melanoma: from mummies to mutations. Melanoma Research 2012;22(2):114-122. Available from:

Crowson AN, Magro CM, Mihm Jr MC. Prognosticators of melanoma, the melanoma report, and the sentinel lymph node. Modern Pathology 2006;19:S71-S87. Available from:

Rashed H, Flatman K, Bamford M, Teo KW, Saldanha G. Breslow density is a novel prognostic feature in cutaneous malignant melanoma. Histopathology 2017;70(2):264-272. Available from:

Marghoob AA, Koenig K, Bittencourt FV, Kopf AW, Bart RS. Breslow thickness and Clark level in melanoma: support for including level in pathology reports and in American Joint Committee on Cancer Staging. Cancer 2000;88(3):589-595.

Massi G, Leboit PE. Criteria for the diagnosis of nevus. Histological Diagnosis of Nevi and Melanoma. 2nd ed. Heidelberg: Springer, 2014:3.

Goldstein AM, Tucker MA. Dysplastic nevi and melanoma. Cancer Epidemiology, Biomarkers and Prevention 2013;22(4):528-532. Available from:

Balu M, Kelly KM, Zachary CB, et al. Distinguishing between benign and malignant melanocytic nevi by in vivo multiphoton microscopy. Cancer Research 2014;74(10):2688-2697. Available from:

Hieken TJ, Hernandez-Irizarry R, Boll JM, Coleman JEJ. Accuracy of diagnostic biopsy for cutaneous melanoma: implications for surgical oncologists. International Journal of Surgical Oncology 2013:196493. Available from:

Troxel DB. Pitfalls in the diagnosis of malignant melanoma: findings of a risk management panel study. The American Journal of Surgical Pathology 2003;27(9):1278-1283. Available from:

Ramos-Vara JA, Miller MA. When tissue antigens and antibodies get along: revisiting the technical aspects of immunohistochemistry – the red, brown and blue technique. Veterinary Pathology 2014;51(1):42-87. Available from:

van der Loos C (2009). User Protocol: practical guide to multiple staining.

Compton LA, Murphy GF, Lian CG. Diagnostic immunohistochemistry in cutaneous neoplasia: an update. Dermatopathology 2015;2(1):15-42. Available from:

Patrascu OM, Costache M, Dumitru AV, et al. Expression of Bcl-2, Melan A and HMB-45 in Dysplastic Nevi. Maedica 2016;11(1):38-43.

Ohsie SJ, Sarantopoulos GP, Cochran AJ, Binder SW. Immunohistochemical characteristics of melanoma. Journal of Cutaneous Pathology 2008;35(5):433-444. Available from:

Prieto VG, Shea CR. Immunohistochemistry of melanocytic proliferations. Archives of Pathology & Laboratory Medicine Online 2011;135:853-859. Available from:

Cockerell CJ, Tschen J, Evans B, et al. The influence of a gene expression signature on the diagnosis and recommended treatment of melanocytic tumors by dermatopathologists. Medicine 2016;95(40):e4887. Available from:

Reddy KK, Rogers GS. Management of dysplastic nevi: the role of complete surgical excision. The Melanoma Letter 2014;32(2).

Nielsen PS, Riber-Hansen R, Steiniche T. Immunohistochemical double stains against Ki67/MART1 and HMB45/MITF: promising diagnostic tools in melanocytic lesions. The American Journal of Dermatopathology 2011;33(4):361-370.