FIG. 4.

Far-Western blot analysis using EREx distinguished the receptor-binding capabilities of the three gC:EPO recombinant molecules. Vero cells were mock transfected (lanes 1 and 2) or transfected with the indicated plasmids. The detergent-solubilized monolayers were immunoprecipitated with α-gC or α-EPO, as indicated. The protein A-bound immune complexes were separated by SDS-PAGE, transferred to a nitrocellulose membrane, and incubated with 5 μg of EREx, a mixture of polyclonal antibodies against the EPO receptor and GST, and a polyclonal antibody conjugated with HRP. Detection was performed using an ECL kit as described by the manufacturer. The mobility of each mature (∗) or premature (○) protein, as well as that of nonspecific protein binding (▵), is indicated.