von Lode Piia, Rosenberg Jaana, Pettersson Kim, Takalo Harri
Department of Biotechnology, University of Turku, Tykistökatu 6A, 6th floor, FIN-20520 Turku, Finland.
Anal Chem. 2003 Jul 1;75(13):3193-201. doi: 10.1021/ac0340051.
New labels and assay techniques are needed to improve the sensitivity and quantitativeness of point-of-care immunotesting while sustaining the rapidity and ease of use of the assays. We synthesized a novel, intrinsically fluorescent nonadentate europium chelate with two chromophores and hydrophilic alpha-galactose side groups. The chelate is highly fluorescent, soluble in water, and provides effective shielding of Eu from water. The performance of the nonadentate chelate was compared with a heptadentate chelate in a dry reagent immunoassay for human chorionic gonadotropin (hCG). After 15-min incubation and washing, time-resolved fluorescence was measured directly from a wet or dried well surface. Contrary to the heptadentate label, the effect of aqueous quenching on the nonadentate label was found to be insignificant, with calculated analytical detection limits (background + 3 SD) of 0.9 and 0.7 IU/L hCG for wet and dry measurements, respectively, and a linear range up to 5000 IU/L. The CVs for the new label were <8% at the cutoff of 25 IU/L and above in both whole blood and plasma. The novel nonadentate label facilitates short turnaround times and simple instrumentation due to the absence of all signal development steps, at the same time retaining an excellent immunoassay performance.
