Time-resolved immunofluorometric assay for thyroxine-binding globulin in serum.
We describe a new "sandwich"-type immunofluorometric assay for thyroxine-binding globulin (TBG) in serum. The assay involves a solid-phase monoclonal antibody immobilised in white microtiter wells, and a soluble biotinylated monoclonal antibody that reacts with the captured TBG molecules. Addition of streptavidin labeled with the europium chelator, BCPDA (4,7-bis(chlorosulfophenyl)-1,10 phenanthroline-2,9-dicarboxylic acid), and excess europium results in the formation of a highly fluorescent product. The fluorescence signal of the final complex is quantitated on the dried solid-phase with a pulsed-laser time-resolved fluorometer. The assay requires a 151-fold sample pre-dilution and a total incubation time of 90 minutes. It has a broad dynamic range of 0-100 mg/L and a minimum detection limit of 0.4 mg/L. The coefficients of variation for within-run and between-run assays averaged 4.5% and 5.4%, respectively. The mean analytical recovery of TBG added to serum was 103%. Results obtained by this method correlated well with those determined by a commercial radioimmunoassay (r = 0.96, n = 112) and by an immunoradiometric procedure (r = 0.95, n = 131).
Tan
Y K
YK
CyberFluor Inc., Toronto, Ontario, Canada.
Khosravi
M J
MJ
Diamandis
E P
EP
D005069:Evaluation Studies as Topic; D015200:Fluoroimmunoassay / Q000379:methods*; D006801:Humans; D013975:Thyroxine-Binding Proteins / Q000032:analysis* / Q000592:standards
D013975:Thyroxine-Binding Proteins