Development of a novel scintillation proximity radioimmunoassay for platelet-activating factor measurement: comparison with bioassay and GC/MS techniques.
A novel, facile and sensitive scintillation proximity radioimmunoassay (SPRIA) for quantitation of PAF has been developed. No separation of antibody bound [3H]PAF from free [3H]PAF is required as the assay employs protein A - coated fluomicrospheres (beads containing scintillant). The assay system was suitable for the quantitation of 0.03 to 2 pmol of 1-hexadecyl-2-acetyl-sn-glycero-3- phosphocholine. The cross-reactivity was high with 1-alkyl-2-acetyl-sn-glycero-3-phosphocholine but was very low with PAF analogs such as 1-alkyl- and 1-acyl-2-lyso-sn-glycero-3-phosphocholine, 1-acyl-2-acetyl-sn-glycero-3-phosphocholine, and 1-alk-1'-enyl-2-acetyl-sn-glycero-3-phosphocholine. The specificity of SPRIA was higher than that of bioassay (platelet degranulation assay). PAF receptor antagonists (L-652,731, WEB2086, and FR900452) at up to 10 nmol per tube had no affect on the SPRIA. These observations indicate that the specificity of the PAF antibody is quite different from that of the platelet receptor. The values obtained using SPRIA for the measurement of PAF produced in polymorphonuclear leukocytes with stimuli are comparable to those obtained by SIM/GC/MS analysis.
Sugatani
J
J
Department of Medical Chemistry, Kansai Medical School, Osaka, Japan.
Lee
D Y
DY
Hughes
K T
KT
Saito
K
K
D000818:Animals; D000918:Antibody Specificity; D000001:Calcimycin / Q000276:immunology; D015550:Cell Degranulation; D008401:Gas Chromatography-Mass Spectrometry; D006801:Humans; D008213:Lymphocyte Activation; D008863:Microspheres; D009504:Neutrophils / Q000032:analysis; D010972:Platelet Activating Factor / Q000032:analysis*; D011817:Rabbits; D011863:Radioimmunoassay / Q000379:methods*; D015203:Reproducibility of Results; D012588:Scintillation Counting / Q000379:methods*; D017640:Silicates; D012824:Silicic Acid; D015019:Yttrium
D010972:Platelet Activating Factor; D017640:Silicates; D012824:Silicic Acid; D000001:Calcimycin; C028301:yttrium silicate; D015019:Yttrium