Melanin fate in the human epidermis: a re-assessment of how best to detect and analyze histologically

Abstract

Melanin is the predominant pigment responsible for skin colour, and is synthesized by the melanocyte in the basal layer of the epidermis and then transferred to surrounding keratinocytes. Despite its optical properties, melanin is barely detectable in unstained sections of human skin. However, identification and localization of melanin is of importance for the study of skin pigmentation in health and disease. Current methods for the histologic quantification of melanin are suboptimal, and are associated with significant risk of misinterpretation. The aim of this study was to re-assess the existing literature, and to develop a more effective histological method of melanin quantification in human skin. Moreover, we confirm that Warthin-Starry (WS) stain provides a much more sensitive and more specific melanin detection method than the common-place Fontana-Masson (FM) stain. For example, WS staining sensitivity allowed the visualization of melanin even in very pale Caucasian skin that was missed by FM or Von Kossa (VK) stains. From our re-assessment of the histologyrelated literature we conclude that so-called ‘melanin dust’ is most likely an artefact of discoloration due to non-specific silver deposition in the stratum corneum. Unlike FM and VK, WS was not associated with this non-specific stratum corneum darkening, misinterpreted previously as ‘degraded’ or so-called ‘dust’ melanin. Finally, WS melanin particle counts were largely similar to manual counts by transmission electron microscopy, in contrast to both FM and VK. Together these findings allow us to propose a new histology/Image J-informed method for the accurate and precise quantification of epidermal melanin in skin.