Time-resolved immunofluorometric assay of alpha-fetoprotein in serum and amniotic fluid, with a novel detection system.
We describe a new "sandwich"-type nonisotopic immunoassay of alpha-fetoprotein (AFP) in serum and amniotic fluid. In the assay, AFP binds to a monoclonal antibody immobilized in a microtiter well and to a polyclonal soluble biotinylated antibody. A fluorescent product is created on the solid-phase after adding streptavidin labeled with a new Eu3+ chelate, 4,7-bis(chlorosulfophenyl)-1,10-phenanthroline-2,9-dicarboxylic acid (BCPDA), and excess Eu3+. The fluorescent complex, monoclonal antibody-AFP-polyclonal antibody-biotin-streptavidin-BCPDA-Eu3+, is quantified by nitrogen laser excitation at 337.1 nm, the emission at 615 nm being monitored in an especially designed gated fluorometer working in a time-resolved mode. This assay of AFP has a broad dynamic range (up to 1 mg/L), and is precise and accurate. The detection limit is approximately 0.1 microgram/L. Results agree well with those obtained by established techniques.
D000653:Amniotic Fluid / Q000032:analysis*; D000911:Antibodies, Monoclonal; D001426:Bacterial Proteins; D001710:Biotin; D005063:Europium; D005260:Female; D005456:Fluorescent Dyes; D006801:Humans; D007118:Immunoassay; D010618:Phenanthrolines; D011247:Pregnancy; D011786:Quality Control; D011863:Radioimmunoassay; D013050:Spectrometry, Fluorescence; D019809:Streptavidin; D000509:alpha-Fetoproteins / Q000032:analysis*
D000911:Antibodies, Monoclonal; D001426:Bacterial Proteins; D005456:Fluorescent Dyes; D010618:Phenanthrolines; D000509:alpha-Fetoproteins; C054206:4,7-bis(chlorosulfophenyl)-1,10-phenanthroline-2,9-dicarboxylic acid; D005063:Europium; D001710:Biotin; D019809:Streptavidin