Development of a sensitive and specific assay system for cholecystokinin tetrapeptide.
Cholecystokinin is a gastrointestinal and neuropeptide which has been implicated in a wide range of physiological and behavioral processes. We have developed a sensitive and specific assay system to measure the various forms of cholecystokinin (CCK) in human plasma. This 3-step system involves i) extraction of CCK fragments from plasma using reverse phase chromatography; ii) separation of peptides by high performance liquid chromatography; and iii) detection and quantification of peptides with a double-antibody radioimmunoassay, using an antibody raised against cholecystokinin tetrapeptide (CCK-4) coupled to thyroglobulin and 125I Bolton-Hunter CCK-4 as tracer. The antibody detects CCK-4, sulfated CCK-8 (CCK-8S) and nonsulfated CCK-8 (CCK-8ns) with equal affinity. The lower limit of detection is 2.7 fmol, with an ED50 of 10.6 +/- 2.2 fmol. Mean CCK-like immunoreactivity (CCK-LI) in the plasma of 12 healthy subjects was determined to be 12.9 +/- 2.1 pM CCK-4 equivalents. Concentrations of each individual peptide in plasma were determined to be 1.0 +/- 0.2 pM, 3.4 +/- 0.8 pM and 1.9 +/- 0.4 pM for CCK-4, CCK-8s and CCK-8ns respectively.
Merani
S
S
Department of Biology, McGill University, Montréal, Québec, Canada.
Palmour
R M
RM
Bradwejn
J
J
Berezowska
I
I
Vaccarino
F J
FJ
Gutkowska
J
J
D000328:Adult; D000818:Animals; D002851:Chromatography, High Pressure Liquid; D005260:Female; D006801:Humans; D008297:Male; D011817:Rabbits; D011863:Radioimmunoassay; D012680:Sensitivity and Specificity; D012844:Sincalide / Q000031:analogs & derivatives / Q000097:blood; D013758:Tetragastrin / Q000097:blood*
C090019:8-sulfocholecystokinin octapeptide; D013758:Tetragastrin; D012844:Sincalide