Fig. 5. Ultrastructural analysis of LGP85-induced large vacuoles. COS cells were transiently transfected with LGP85 and, after 36 hours of transfection, were processed for immunoelectron microscopy. Ultra-thin sections were either single-labeled with anti-LGP85, followed by anti-rabbit IgG-10 nm gold (A,B) or double-labeled with anti-LGP85, and anti-LAMP-2 (C, arrowheads), or anti-LGP85 and anti-LBPA (D,E, arrowheads), followed by anti-rabbit IgG-10 nm gold and anti-mouse IgG-5 nm gold. Arrowheads in A show the double limiting membrane structure of LGP85-induced large vacuoles, and a higher magnification image is shown in B. Panel D shows a portion of the limiting membrane of one large LGP85-containing vacuole, with the lumen of the vacuole occupying the upper half of the panel. Note that some LBPA labeling is seen on internal membranes closely associated with the limiting membrane of the vacuole (D, arrows). Bars, 200 nm.