Clathrin-coated vesicles budding from endosomes are deficient in VSG. Cryosections were labeled with anti-VSG antibodies and PAG-6 (A–C) or double labeled with anti-clathrin and anti-VSG antibodies and PAG-6, PAG-10, or PAG-15 as indicated (D–F). (A) An EC with numerous clathrin-coated buds has been sectioned perpendicularly in the upper part and in plane in the lower part. Whereas the lumen of the cisterna is strongly labeled for VSG, the buds are nearly VSG free. (B) The buds (arrows) as well as an adjacent region of the cisterna (arrowheads) are clathrin-coated but free of VSG. The latter region may indicate an incipient clathrin-coated bud. (C) A very thin cryosection with a single gold grain inside the terminal bud (arrow) at the rim of the cisterna. (D–F) Budding structures on cisternae marked by arrows are positive for clathrin and negative for VSG, whereas the reverse pattern is observed for their sheet-like core region. Adjacent, free clathrin-coated vesicles (E and F, arrowheads) are also free of VSG. SC, surface coat. Bars (A, B, E, and F), 125 nm; 250 nm (C); and 500 nm (D).