title: Method of melanin bleaching in MIB1-Ki67 immunostaining of pigmented lesions: A quantitative evaluation in malignant melanomas.
abstract: The effect of melanin bleaching on the immunoreactivity of the MIB1-Ki67 antigen in pigmented melanocytic lesions was investigated. Eight paired non-pigmented and heavily pigmented malignant melanomas (6 primary melanomas and 2 secondary melanomas) were selected. Avidin-biotin immunoperoxidase complex (ABC) and microwave antigen retrieval were used in immunostaining. Sections were incubated with 10% H2O2 for 24h before immunostaining with primary antibody MIB1, or after the completion of immunostaining. Non-bleached controls were obtained by conducting the identical staining but omitting the bleaching procedure. In all heavily pigmented lesions bleached by 10% H2O2 before or after immunostaining, the melanin was bleached effectively and MIB1-positively stained cells were clearly seen. Cell counting in the non-pigmented group found that there were no significant differences in the percentage of MIB1-positive melanoma cells (%MIB1) between non-bleached controls and those sections which had been bleached by 10% H2O2 either before or after the immunostaining. The results suggest that hydrogen peroxide can effectively bleach melanin in pigmented melanocytic lesions without significantly affecting MIB1-Ki67 immunolabelling.
authors.affiliation: Department of Pathology, University of Sydney, NSW, Australia.
authors.forename: L X
authors.initials: LX
authors.lastname: Li
authors.forename: K A
authors.initials: KA
authors.lastname: Crotty
authors.forename: J J
authors.initials: JJ
authors.lastname: Kril
authors.forename: A A
authors.initials: AA
authors.lastname: Palmer
authors.forename: S W
authors.initials: SW
authors.lastname: McCarthy
mesh_terms: D000911:Antibodies, Monoclonal; D000918:Antibody Specificity; D002452:Cell Count; D016591:Histocytological Preparation Techniques; D006801:Humans; D006861:Hydrogen Peroxide; D007150:Immunohistochemistry / Q000379:methods; D019394:Ki-67 Antigen / Q000737:chemistry*; D008543:Melanins / Q000737:chemistry*; D008544:Melanocytes / Q000737:chemistry* / Q000473:pathology; D008545:Melanoma / Q000737:chemistry* / Q000473:pathology; D016877:Oxidants; D012878:Skin Neoplasms / Q000737:chemistry* / Q000473:pathology
chemical_list: D000911:Antibodies, Monoclonal; D019394:Ki-67 Antigen; D008543:Melanins; D016877:Oxidants; D006861:Hydrogen Peroxide