A rapid, high capacity assay for basic fibroblast growth factor binding.
A rapid, high capacity assay for the binding of basic fibroblast growth factor has been developed. Rat lung tissue was selected as the optimum source of membranes and 2M sodium chloride used to remove endogenous growth factor. The assay has been adapted to the Millipore Multi-Screen system so that it can be run in 96-well format with a volume of 300 microliters. The assay has been validated through the demonstration of inhibition by standard inhibitors such as suramin and protamine sulfate. The assay has proven useful for the screening of random compounds as well as the more detailed examination of suspected inhibitors. By running compounds in the presence and absence of a mid-range concentration of unlabeled bFGF, an estimate of the proportion of the inhibition due to high affinity binding can be obtained. Suramin and protamine sulfate show no selectivity and inhibit high affinity binding and overall binding with similar potencies. Another inhibitor, dimercaptothiadiazole, is more potent against high affinity binding.
Herblin
W F
WF
DuPont Merck Pharmaceutical Company, Wilmington, DE 19880-0400, USA.
Barbera
F A
FA
D000818:Animals; D016222:Fibroblast Growth Factor 2 / Q000037:antagonists & inhibitors / Q000378:metabolism*; D066298:In Vitro Techniques; D008168:Lung / Q000378:metabolism; D011479:Protamines / Q000494:pharmacology; D011485:Protein Binding; D051381:Rats; D017468:Receptors, Fibroblast Growth Factor / Q000378:metabolism; D013498:Suramin / Q000494:pharmacology; D013830:Thiadiazoles / Q000494:pharmacology
D011479:Protamines; D017468:Receptors, Fibroblast Growth Factor; D013830:Thiadiazoles; D016222:Fibroblast Growth Factor 2; C012291:2,5-dimercapto-1,3,4-thiadiazole; D013498:Suramin