Bleaching Melanin in Formalin-fixed Paraffin Wax-embedded Pigmented Melanomas Using Visible Light

Introduction: Melanin in histological sections can hamper the evaluation of cellular details in pigmented melanocytic tumours. In order to remove melanin from formalin-fixed and paraffin wax-embedded (FFPE) sections, chemicals are generally used. Since melanin absorbs light, our aim was to test whether visible light can bleach pigmented melanoma FFPE sections maintaining cell morphology. Materials and Methods: Serial 4 mm FFPE sections of an equine, a canine and a porcine cutaneous pigmented melanoma and a feline diffuse iris pigmented melanoma were dewaxed, rehydrated and dipped in Tris-buffered saline. For each sample, five sections were irradiated with a visible light-emitting LED spotlight for 1, 2, 3, 4 and 5 days, respectively, while a control section was not irradiated. During the procedure, sections were maintained at the same temperature. All sections were stained concurrently with haematoxylin and eosin. Pigment degradation and cell morphology maintenance were evaluated comparing bleached and control sections. Results: Complete bleaching was obtained after 1 day of treatment of the feline and swine melanomas, while 2 and 3 days were required for the canine and equine melanomas, respectively. Following phototreatment the cell morphology was well maintained and bleached cells revealed excellent cytoplasmic and nuclear details. Discussion: Visible light can bleach FFPE specimens of pigmented melanoma without affecting the cell morphology. Although results were very good in terms of pigment degradation and cell morphology preservation, a limit of this method is the long time required to obtain complete depigmentation. Exploiting different variables to reduce the bleaching time is our future goal.