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Figure 1


Fig. 1. siRNA-mediated depletion of clathrin heavy chain blocks endocytosis of TfnR. (A) HeLa cells were transfected with siRNA oligonucleotides targeting clathrin heavy chain (CHC). Whole-cell extracts were prepared and subjected to immunoblotting using anti-CHC or actin antibodies. (B,C) Control siRNA (B) or CHC siRNA (C) cells were incubated with Alexa-Fluor-546-labeled Tfn before being fixed and subjected to indirect immunofluorescence using anti-CHC antibody. (D) HA-tagged dynamin(PH*)-transfected cells were incubated with Alexa-Fluor-546-labelled Tfn, then fixed and subjected to indirect immunofluorescence using anti-HA antibody. (E) CHC siRNA cells were incubated with Alexa-Fluor-488-labeled Tfn, then fixed and subjected to indirect immunofluorescence using anti-Tfn receptor antibody. All samples were analyzed by confocal microscopy. Arrows indicate depleted cells.





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