Fig. 2. Relative protein expression levels of HA-GLUT4 constructs. Isolated rat adipose cells were transfected with 1.25 µg/ml plasmid DNA (12.5 µg/ml for the 37 mutant) and cultured for 24 hours. After harvesting, total cellular membranes were prepared and the expression of HA-GLUT4 was determined by western blot using anti-HA antibodies as described in Materials and Methods (inset). The HA-GLUT4 proteins migrated according to their expected molecular masses in SDS-PAGE and appeared as single bands on western blots. Results are the means±s.e.m. of at least three independent experiments.