Characterization of a scintillation proximity assay to detect modulators of transforming growth factor alpha (TGF alpha) binding.
A scintillation proximity assay (SPA) for transforming growth factor alpha (TGF alpha) using SPA beads coated with A431 membranes has been studied. Binding of TGF alpha to the beads was characteristic of a receptor interaction. A class of high-affinity receptors for [125I]-TGF alpha (Kd = 0.10-0.26 nM) was detected by competition studies between [125I]TGF alpha and cold TGF alpha and by analysis of association and dissociation rate constants. An antibody to the epidermal growth factor receptor (clone 528) inhibited binding of [125I]TGF alpha (IC50 = 0.20 micrograms/ml), but an anti-TGF alpha antibody (clone 134A-2B3) (less than 25 micrograms/ml) did not block binding. Suramin inhibited [125I]-TGF alpha binding (IC50 = 0.20 mM). The ether lipids 1-O-hexadecyl-2-O-methyl-sn-glycero-3-phosphocholine, 1-O-octadecyl-2-O-methyl-sn-glycero-3-phosphocholine, and rac-lyso-platelet activating factor inhibited TGF alpha binding (IC50 values of 49, 69, and 57 microM, respectively). SPA is a convenient method for identifying agents that may act by interfering with TGF alpha binding.
Hoffman
R
R
Xenova Ltd., Slough, England.
Cameron
L
L
D000911:Antibodies, Monoclonal; D001667:Binding, Competitive; D002460:Cell Line; D066246:ErbB Receptors / Q000037:antagonists & inhibitors; D007700:Kinetics; D012588:Scintillation Counting / Q000379:methods*; D013498:Suramin / Q000494:pharmacology; D016211:Transforming Growth Factor alpha / Q000032:analysis*
D000911:Antibodies, Monoclonal; D016211:Transforming Growth Factor alpha; D013498:Suramin; D066246:ErbB Receptors