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Fig. 4. PGI/AMF endocytosis is required for its association with cell surface FN fibrils. NIH-3T3 cells were uninfected (A,B) or infected for 48 hours with adenoviruses expressing wild-type dynamin-1 (dynWT) (C,D), the dominant negative dynamin-1 K44A mutant (dynK44A) (E,F) and the dominant negative clathrin hub (Cla-Hub) (G,H). The cells were then incubated with 25 µg ml–1 of PGI/AMF-568 for 60 minutes (A,C,E,G) and fixed with 3% paraformaldehyde. Infection rates were from 50-80% and in the selected fields, all cells were infected. Uninfected, dynWT and dynK44A infected cells were labeling with mouse anti-HA antibody and goat Alexa-488 anti-mouse secondary antibody to identify infected cells (B,D,F) and clathrin hub infected cells were identified by labeling with mouse anti-T7 antibody followed by labeling with goat Alexa-488 anti-mouse secondary antibody (H). Bar, 20 µm.
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