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Fig. 6. Rabip4 translocates to the plasma membrane in response to insulin. (A) Plasma membrane sheets were prepared from 3T3-L1 adipocytes, untreated or treated with 100 nM insulin, for 20 minutes. Rabip4 and Glut 4 were detected by immunofluorescence using rabbit anti-Rabip4 antibodies and goat anti-Glut 4 antibodies followed by incubation with FITC-coupled anti-rabbit antibodies and Texas-Red-coupled anti-goat antibodies, respectively. Bar, 10 µm. (B) Quantification of fluorescence levels from A, performed using MetaMorph software as described in the Materials and Methods. (C) 3T3-L1 adipocytes were untreated or treated with 100 nM insulin for 20 minutes. Plasma membranes (PM), intracellular microsomes (IM), and cytosol (Cyt) were then prepared and 40 µg proteins from each fraction were separated by SDS-PAGE and transferred to PVDF membrane. Antibodies against Rabip4, Glut4 and Rab4 were then used to probe the blot.
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