Cationic lipid reagents activate and down-regulate the insulin receptor. Upper panel: Cho.hIR cells were incubated with LF2000 (Lip) or FuGENE (Fug) as described in the methods section or were treated with insulin (10mins) or medium (C). Cell lysates were blotted for phosphotyrosine and the position 97kDa molecular weight marker (which coincides with the insulin receptor beta chain) is indicated by the arrow. Middle panel: Cho.hIR cells were treated as above except that treatment with LF2000 and FuGENE lasted 24 hours and blotted for phosphotyrosine. Lysates labelled "I" are from cells incubated for 24 hours as control cells and subsequently stimulated with insulin for 10 minutes to confirm that cells are otherwise responsive to insulin after the incubation period. The position of the 97kDa marker is indicated by the arrow. Lower panel: The same cells as in the middle panel were blotted for the presence of the insulin receptor, which co-migrates with the 97kDa marker, indicated by the arrow. The blots are from a representative experiment repeated three times.
Pramfalk et al. BMC Cell Biology 2004 5:7 doi:10.1186/1471-2121-5-7