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Improved Immunohistochemistry of Mouse Eye Sections Using Davidson's Fixative and Melanin Bleaching
Improved Immunohistochemistry of Mouse Eye Sections Using Davidson's Fixative and Melanin Bleaching
Anne Longakit
(Department of Medical Genetics, University of British Columbia, Vancouver, BC, CanadaLife Sciences Institute, University of British Columbia, Vancouver, BC, Canada,)
Chloe Hess
(Department of Medical Genetics, University of British Columbia, Vancouver, BC, CanadaLife Sciences Institute, University of British Columbia, Vancouver, BC, Canada,)
Christina Zhang
(Department of Medical Genetics, University of British Columbia, Vancouver, BC, CanadaLife Sciences Institute, University of British Columbia, Vancouver, BC, Canada,)
Catherine Van Raamsdonk
(Department of Medical Genetics, University of British Columbia, Vancouver, BC, CanadaLife Sciences Institute, University of British Columbia, Vancouver, BC, Canada,)
Immunohistochemistry (IHC) and immunofluorescence (IF) are fundamental molecular biology techniques to assess protein expression. However, the melanin present normally in the eye in the uveal tract (choroid, iris, and ciliary body) and the retinal pigment epithelium (RPE) poses a significant challenge for IHC and IF. This is because melanin interferes with both chromogenic and fluorescent detection methods. Additionally, formalin fixation, which is commonly used for IHC, can result in shrinkage and loss of cellular detail in the eye. This protocol provides an optimized approach using Davidson’s fixative with a hydrogen peroxide bleaching step to eliminate melanin interference in the mouse eye, improving the quality and interpretability of IHC analyses of the uveal tract and RPE. It is particularly useful for the analysis of uveal melanoma.
Anne Longakit, Chloe Hess, Christina Zhang, et al. (2025). "Improved Immunohistochemistry of Mouse Eye Sections Using Davidson's Fixative and Melanin Bleaching". Bio-Protocol. https://doi.org/10.21769/bioprotoc.5510