Determination of prostaglandin metabolites in biological samples by competitive time-resolved fluoroimmunoassay.
We describe a time-resolved fluoroimmunoassay method for the determination of 13,14-dihydro-15-ketoprostaglandin E2 (PGEM) and 13,14-dihydro-15-ketoprostaglandin F2 alpha (PGFM) in femtomolar ranges. Polyclonal antibodies were raised in rabbits and the antigen was labelled with europium by the avidin-biotin technique. A second antibody, directed to rabbit IgG, was coated onto the solid phase. The IgG-PG-europium complex was bound by the second antibody, giving a rapid and complete separation of antibody-bound and free antigen. The detection limit was 0.2 pg/assay for PGFM and 1.2 pg/assay for PGEM. The intra-assay CV ranged from 4.6% to 9.3% and the interassay CV from 6.7% to 11.4%. A good correlation was obtained between the results from TR-FIA and RIA when the method was applied to the investigation of tissues from breast cancer. We conclude that the TR-FIA is more sensitive and much faster than the RIA and avoids the use of radioactive material.
Lüke
F J
FJ
Department of Obstetrics and Gynaecology, University of Münster, Germany.
Schlegel
W
W
D001426:Bacterial Proteins; D001667:Binding, Competitive; D001710:Biotin; D001943:Breast Neoplasms / Q000378:metabolism; D015237:Dinoprost / Q000031:analogs & derivatives / Q000032:analysis; D015232:Dinoprostone / Q000031:analogs & derivatives / Q000032:analysis; D005063:Europium; D015200:Fluoroimmunoassay / Q000379:methods*; D006801:Humans; D011453:Prostaglandins / Q000378:metabolism*; D011863:Radioimmunoassay; D012015:Reference Standards; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D019809:Streptavidin
D001426:Bacterial Proteins; D011453:Prostaglandins; C008228:15-keto-13,14-dihydroprostaglandin F2alpha; C025087:15-keto-13,14-dihydroprostaglandin E2; D005063:Europium; D001710:Biotin; D019809:Streptavidin; D015237:Dinoprost; D015232:Dinoprostone