Method of Melanin Bleaching in MIB1-Ki67 Immunostaining of Pigmented Lesions: A Quantitative Evaluation in Malignant Melanomas |
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Authors: | Ling-Xi Lawrence Li Kerry A. Crotty Jillian J. Kril Allan A. Palmer Stanley W. McCarthy |
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Affiliation: | (1) Department of Pathology, University of Sydney, NSW 2006, Australia;(2) Department of Anatomical Pathology, Royal Prince Alfred Hospital, Camperdown, NSW 2050, Australia;(3) Centre for Education and Research on Ageing (CERA), Concord Hospital, Concord, NSW, 2139, Australia |
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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. |
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