Preparation and Characterization of a Fully Active Biotinylated Probe of Cholecystokinin |
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| Authors: | R. S. Izzo C. Pellecchia M. I. Weinstein A. J. Massimillo |
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| Affiliation: | Department of Medicine , Gastroenterology Division Nassau County Medical Center , East Meadow, New York |
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| Abstract: | In this study we describe the synthesis and purification of biotinylated cholecystokinin-8 (Bio-CCK-8) and characterize its use as a probe for the pancreatic cholecystokinin receptor. CCK-8 (0.1 umoles) was reacted with either radiolabeled d-[8,9-3H]biotin succinimide ester (0.5 umoles) or N-hydroxysuccinimidyl-biotin in dimethylformamide and triethylamine, and purified by anion exchange chromatography. Concentrations of Bio-CCK-8 and CCK-8 needed for half-maximal inhibition of [l25]I-CCK-8 binding to pancreatic membranes were the same (1.0 and 1.3 nM). Bio-CCK-8 retained full biological activity as determined by stimulation of pancreatic protein secretion from rats, and the biotin group bound to CCK-8 retained its high sensitivity for avidin. |
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