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In the course of our study concerning gastrin and cholecystokinin (CCK) receptors, we have synthesized and characterized a new labeled gastrin ligand, 125I-BH-[Leu15]-gastrin-(5-17) [(3-[125I]iodo-4-hydroxyphenyl)-propionyl-[Leu15]-gastrin-(5-17)]. Binding of 125I-BH-[Leu15]-gastrin-(5-17) to isolated canine fundic mucosal cells was specific, saturable and of high affinity. 125I-BH-[Leu15]-gastrin- (5-17) and 125I-BH-CCK-8[(3-[125I]iodo-4-hydroxyphenyl)-propionyl-CCK-8] interact with isolated canine fundic mucosal cells with small differences in maximal binding capacities and affinities, 3800 +/- 900 binding sites/cell (Kd = 0.52 +/- 0.23 nM) and 6200 +/- 1100 binding sites/cell (Kd = 0.31 +/- 0.18 nM), respectively. The relative order of potencies for gastrin and CCK analogs in displacing 125I-BH-[Leu15]-gastrin-(5-17) binding correlated well with those obtained using 125I-BH-CCK-8. Selective CCK/gastrin antagonists L-364,718 (MK-329) and L-365,260 also inhibited 125I-BH-[Leu15]-gastrin-(5-17) binding. These results indicate that 125I-BH-[Leu15]-gastrin-(5-17) binds to gastrin receptors in isolated canine fundic mucosal cells. We have also characterized 125I-BH-[Leu15]-gastrin-(5-17) binding to the human Jurkat lymphoblastic cell line (Jurkat cells) known to express the CCK-B/gastrin receptor. Saturation experiments have shown that both 125I-BH-[Leu15]-gastrin-(5-17) and 125I-BH-CCK-8 interact with a single class of high-affinity binding sites in the Jurkat cell line.(ABSTRACT TRUNCATED AT 250 WORDS)