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ARTICLE |
Reduction of Lipofuscin-like Autofluorescence in Fluorescently Labeled Tissue
Stephen A. Schnella, William A. Stainesb, and Martin W. Wessendorfaa Department of Cell Biology and Neuroanatomy, University of Minnesota, Minneapolis, Minnesota
b Department of Cellular and Molecular Medicine, University of Ottawa, Ottawa, Ontario, Canada
Correspondence to: Stephen A. Schnell, 4-102 Owre Hall, Dept. Cell Biology and Neuroanatomy, 321 Church Street SE, U. of Minnesota, Minneapolis, MN 55455.
The fluorescent pigment lipofuscin accumulates with age in the cytoplasm of cells of the CNS. Because of its broad excitation and emission spectra, the presence of lipofuscin-like autofluorescence complicates the use of fluorescence microscopy (e.g., fluorescent retrograde tract tracing and fluorescence immunocytochemistry). In this study we examined several chemical treatments of tissue sections for their ability to reduce or eliminate lipofuscin-like autofluorescence without adversely affecting other fluorescent labels. We found that 1–10 mM CuSO4 in 50 mM ammonium acetate buffer (pH 5) or 1% Sudan Black B (SB) in 70% ethanol reduced or eliminated lipofuscin autofluorescence in sections of monkey, human, or rat neural tissue. These treatments also slightly reduced the intensity of immunofluorescent labeling and fluorescent retrograde tract tracers. However, the reduction of these fluorophores was far less dramatic than that for the lipofuscin-like compound. We conclude that treatment of tissue with CuSO4 or SB provides a reasonable compromise between reduction of lipofuscin-like fluorescence and maintenance of specific fluorescent labels. (J Histochem Cytochem 47:719–730, 1999)
Key Words: copper sulfate, Sudan Black B, monkey, human, primate, rat, immunohistochemistry, immunocytochemistry, retrograde axonal tract tracing
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