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Fig. 4. Overexpression of Rab7T22N accumulates large autophagosomes. (A) The levels of overexpressed EGFP proteins were detected by western blot using an anti-EGFP antibody as indicated under Materials and Methods. (B) CHO cells expressing EGFP-Rab7 wt, the mutants Q67L or T22N, or the vector alone (EGFP) were incubated at 37°C for 2 hours in EBSS medium (starved cells). Following this incubation period, cells were incubated with 0.05 mM monodansylcadaverine (MDC) for 10 minutes at 37°C. Intracellular MDC was measured by fluorescence photometry as indicated under Materials and Methods. The data represent the mean ± s.e.m. of five independent experiments. (C,D) The number of MDC-labeled vacuoles per cell (C) and the size (D) were quantified by the Methamorph software, using the integrated morphometry analysis. Data represent the mean±s.e.m. (n=50 cells). **P<0.01.
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