Time-resolved fluoroimmunoassay for pituitary adenylate cyclase activating polypeptide 27 (PACAP27) using europium (III) ion chelate labeled streptavidin-biotin complex.
Pituitary adenylate cyclase activating polypeptide (PACAP) is a novel peptide hormone and has a variety of biological action. In studies of the physiological behaviour of endogenous PACAP, the determination of PACAP levels in biological materials require a highly sensitive and specific method. Therefore, we developed a sensitive time-resolved fluoroimmunoassay (TR-FIA) for PACAP27 which is the biologically important fragment of PACAP. Accordingly, we developed TR-FIA using a biotinylated PACAP27 (b-PACAP27) as a tracer and europium (III) chelate labeled streptavidin-biotinylated bovine serum albumin complex as a detection of biotin on solid phase. A measurable range of PACAP27 was 7.8-1000 pg m1-1 by the proposed TR-FIA. For measurement of biological samples, the samples were purified to eliminate substances which interfered with the TR-FIA. The mean recovery of PACAP27 using commercially reversed phase column was 74.7% (n = 12). The various tissues, extracts and plasma concentrations of rat could be measured by the proposed TR-FIA.
Ito
K
K
School of Pharmaceutical Sciences, Showa University, Tokyo, Japan.
Goto
T
T
Tsuji
A
A
Maeda
M
M
D000818:Animals; D001426:Bacterial Proteins / Q000737:chemistry; D001710:Biotin / Q000737:chemistry; D002614:Chelating Agents; D003429:Cross Reactions; D005063:Europium; D005260:Female; D015200:Fluoroimmunoassay / Q000379:methods*; D016014:Linear Models; D008297:Male; D009479:Neuropeptides / Q000032:analysis*; D018377:Neurotransmitter Agents / Q000032:analysis*; D051219:Pituitary Adenylate Cyclase-Activating Polypeptide; D051381:Rats; D017207:Rats, Sprague-Dawley; D012015:Reference Standards; D012680:Sensitivity and Specificity; D012710:Serum Albumin, Bovine / Q000737:chemistry*; D019809:Streptavidin; D013997:Time Factors
C495403:Adcyap1 protein, rat; D001426:Bacterial Proteins; D002614:Chelating Agents; D009479:Neuropeptides; D018377:Neurotransmitter Agents; D051219:Pituitary Adenylate Cyclase-Activating Polypeptide; D012710:Serum Albumin, Bovine; D005063:Europium; D001710:Biotin; D019809:Streptavidin