Figure 3.

Localization of Rifyfilin-GFP by electron microscopy. Ultrathin cryosections of transfected HeLa cells were single immunogold labeled with anti-GFP and protein A Gold conjugate (PAG10, 10 nm) (A) and double immunogold labeled for TfR (PAG15, 15 nm) and GFP (PAG10) (B) or for CD63 (PAG15) and GFP (PAG10) (C). Cells overexpressing Rffl-GFP display a tubulo-vesicular network of membranes in close apposition to vacuolar endosomes. Rffl-GFP accumulates in these tubulo-vesicular membranes (A). The TfR accumulates in tubulovesicular structures together with Rffl-GFP (B). CD63 is detected in late endosomes/lysosomes that are often close to the Rffl-positive tubules (C). Arrows: 10-nm gold particles; arrowheads: 15-nm gold particles; stars: late endosomes/lysosomes. Bars, 200 nm.