Measurement of Cholecystokinin

Early measurements of cholecystokinin (CCK) were based on the abilities of blood or blood extracts to stimulate gallbladder contraction or pancreatic exocrine secretion. These biological assays were performed in laboratory animals such as dogs, pigs or rodents. The usefulness of these assays was limited because methods were cumbersome and interpretation of results was complicated by confounding problems introduced by other hormones or neural influences. Invention of radioimmunoassay revolutionized the study of endocrinology and most hormones can now be measured by this method. However, the radioimmunoassay of CCK was particularly difficult for several reasons (9). First, multiple molecular forms of CCK ranging from CCK-4 to CCK-83 have been found in blood. Although it appears as though CCK-58 is the major circulating form in the species examined, other forms may contribute to CCK activity. Second, CCK is structurally similar to gastrin. Both CCK and gastrin have an identical pentapeptide carboxyl terminus. Therefore, to avoid crossreactivity, assays must be able to detect the regions of CCK that are distinct from gastrin. Third, CCK circulates in the blood at concentrations that are much lower (10-100 fold) than gastrin. Fourth, large molecular forms of CCK (e.g., CCK-58) that can be used as standards in assays are not routinely available. Fifth, isotope labeling of CCK can be difficult. Sixth, CCK has posttranslationally modified amino acids (e.g., sulfated tyrosine) that are important for activity. CCK-8 is fully active and more potent than shorter forms of CCK. Full activity requires that (1) the hormone be sulfated in position seven (from the carboxyl terminus) and (2) methionine at position 3 remain in the reduced (non-oxidized) state. This multitude of potential problems delayed the development of reproducible, sensitive, and specific RIAs for CCK.