TrueBlack® Lipofuscin Autofluorescence Quencher, 20X in DMF, is a novel reagent that quenches lipofuscin autofluorescence in tissue sections for immunofluorescence staining.
TrueBlack® effectively eliminates lipofuscin autofluorescence in tissues like human and mouse brain and retina (see Figs. 1-3 and TrueBlack® References). Lipofuscin consists of highly autofluorescent granules of oxidized proteins and lipids that build up in the lysosomes of aging cells. Lipofuscin granules fluoresce brightly in all channels used for fluorescence microscopy, and accumulate in a variety of cell and tissue types with age. Consequently, imaging of specific immunofluorescence signal in certain adult human tissues or aged animal tissues can be virtually impossible unless methods are employed to quench or mask lipofuscin fluorescence.
Traditionally, Sudan Black B has been used to quench lipofuscin autofluorescence by incubating tissue sections with the dye after immunofluorescence staining. However, while it masks the autofluorescence from lipofuscin, Sudan Black B also introduces uniform non-specific background fluorescence in the red and far-red channels (Fig. 1), limiting the use of fluorescent dyes in those wavelengths. Now Biotium has developed TrueBlack® as a superior alternative to Sudan Black B. TrueBlack® quenches lipofuscin fluorescence with far less increase in red/far-red background fluorescence (Fig. 1). TrueBlack® treatment can be performed before or after immunostaining. It is rapid, simple, and has minimal effect on signal from fluorescent antibodies or nuclear counterstains, thus preserving the signal-to-noise ratio of the immunostaining (Figs. 2-3).
TrueBlack® also can reduce autofluorescence from other sources, such as collagen, elastin, red blood cells, and general background fluorescence. It is not as effective at quenching these sources of autofluorescence as it is for lipofuscin, but it can improve background in human and non-human tissue types (Figs. 4-5). TrueBlack® also has been used to quench background fluorescence on polycarbonate filters used as cell supports for imaging cytometry.
TrueBlack® quenches lipofuscin with less background than Sudan Black B
TrueBlack® treatment can be performed before or after immunofluorescence staining
TrueBlack® can reduce autofluorescence from sources other than lipofuscin