High-performance assays of small molecules: enhanced sensitivity, rapidity, and convenience demonstrated with a noncompetitive immunometric anti-immune complex assay system for digoxin.
We report the first clinical application of a noncompetitive immunometric assay system that provides advantages for the rapid and robust assay of small molecules similar to those realized for larger molecules with two-site immunometric assays. This anti-immune complex assay is based on the interaction of a receptor such as a primary antibody with its ligand, such that new binding sites, recognizable by a secondary antibody, are formed. In this report the system is applied to the measurement of digoxin in serum. Utilizing an anti-complex antibody that recognizes a digoxin-bound primary antibody with affinity > 2000-fold over its binding to the primary antibody alone, we show that this anti-complex assay system provides a high-performance assay for serum samples, being conveniently simple (immobilized primary antibody binds digoxin and then labeled secondary antibody so that when excess unbound label is washed away the immunometric readout reflects the digoxin concentration), rapid (incubation time 1-10 min), sensitive (detection limit 30 ng/L), precise (3-4% within-run CV, 1-8% total CV), and free from interference from digoxin-like immunoreactive factors.
Department of Clinical Biochemistry, Royal Victoria Infirmary, Newcastle upon Tyne, UK.
C H
CH
Self
J L
JL
Dessi
L A
LA
Winger
D000888:Antibodies, Anti-Idiotypic; D000911:Antibodies, Monoclonal; D000918:Antibody Specificity; D000936:Antigen-Antibody Complex / Q000276:immunology; D001667:Binding, Competitive; D004077:Digoxin / Q000097:blood* / Q000276:immunology; D005260:Female; D006801:Humans; D007118:Immunoassay / Q000379:methods* / Q000706:statistics & numerical data; D008970:Molecular Weight; D011247:Pregnancy; D011786:Quality Control; D051437:Renal Insufficiency / Q000097:blood; D012680:Sensitivity and Specificity; D013997:Time Factors
D000888:Antibodies, Anti-Idiotypic; D000911:Antibodies, Monoclonal; D000936:Antigen-Antibody Complex; D004077:Digoxin