Fig. 6

Identification of Kit as the antigen for scFv A1. a Immunoprecipitation of HMC-1 plasma membrane proteins with scFv A1 identifed a unique band of approximately 145 kDa (arrow). Analogous immunoprecipitations using scFvs C1 (anti-mast cell scFv) and E4 (scFv with irrelevant binding) are shown for comparison. Excision and sequencing demonstrated that this was Kit. b Western blotting of protein immunoprecipitated by scFv A1 demonstrated positive staining using an anti-Kit antibody, with a doublet of the expected size for Kit (145/120 kDa). c Flow cytometry histograms demonstrating that following incubation of HMC-1 with SCF (100 ng/ml) for 15 min, binding of scFv A1 was markedly reduced compared to control, in keeping with SCF-induced Kit internalisation