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Fig. 5. Effect of Alix depletion on transferrin endocytosis and recycling. HeLa cells were treated with control RNA or with siRNA against Alix as in Fig. 1B. For the endocytosis experiments (A), cells were incubated with Ru-tag-Tf for different times at 37°C as described in the Materials and Methods, and the surface-bound Ru-tag-Tf was reduced with MESNA. The amount of cell-associated Tf was measured using an ORIGEN analyser, and endocytosed Tf is presented as a percentage of total cell-associated Tf. Error bars represent the s.e.m. of three independent experiments performed in duplicate. (B) Cells were incubated with Ru-tag-Tf for 10 minutes and then washed and chased at 37°C for 5, 10 or 15 minutes before MESNA treatment. Error bars represent the s.e.m. of three independent experiments performed in duplicate.
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