Time-resolved luminescent immunoassay

Following this study, Korovin and coworkers tested cryptands 23b-f which are used in time-resolved luminescent immunoassays (Mathis, 1993) for the sensitization of Ybm luminescence (Korovin et al., 2002b). From the lifetimes determined in both water and deuterated water, one calculates that the hydration number varies from 2 (23b), to 1.5 (23a, 23e), and finally to 1 (23c, 23d, 23f). Quantum yields were not determined, but luminescence intensities relative to the cryptate with 23a (in water, at room temperature) point to cryptands 23c and 23d being the best sensitizers of the Yb111 luminescence with a seven-fold enhancement, while cryptates with 23e and 23b are only 1.5- to 1.8-times more luminescent. 

Hagan AK, Zuchner T. Lanthanide-Based Time-Resolved Luminescence Immunoassays. Anal Bioanal Chem 2011 400 2847-2864. 

Hagan A. K., T. Zuchner. Lanthanide-based Time-resolved Luminescence Immunoassays, Anal. Bioanal. Chem., 400, 2847-2864 (2011). 

Charbonniere LJ, Hildebrandt N, Ziessel RF, Lohmannsroben HG (2006) Lanthanides to quantum dots resonance energy transfer in time-resolved fluoro-immunoassays and luminescence microscopy. J Am Chem Soc 128 12800-12809... 

LANTHANIDE CHELATES AS LUMINESCENT LABELS IN BIOMEDICAL ANALYSES 195 6. Time-resolved flnorometric immunoassay... 

Different variations of time-resolved luminescence assays were patented in 1982 by Wieder [1], in 1983 by Soini and HemmUa [2], and in 1999 by Diamandis [3]. The Finnish company Wallac first commercialized the principle and introduced an assay reader for dissociation-enhanced lanthanide fluorescence immunoassay (DELFIA) in the beginning of the 1980s. The first (1982) DELFIA-based bioassay for diagnostic market was for Rubella antibodies, and it was the first sensitive nonradioisotope innnunoassay marking the beginning of a new era [4]. 

The application of the Horrock and Sudnick equation to the terbium complex of ligand 6, reveals that only one water molecule is still coordinated to the complexed cation. Consequently, the lifetime (t = 1.5 sec) and the luminescence quantum yield (< > = 0.2) of this complex are very high and make it attractive for applications as luminescent probes in time-resolved fluoro-immunoassays (TR-FIA). " In order to lurther improve the luminescence properties of these ligands by increasing their molar absorbitivities (e) we have synthesized calix[4]arenes (e.g. 7) with four bipyridines or phenantrolines at the lower rim. A significant increase in the value of e of these derivatives has been... 

Figure 19-16 shows how Eu3+ can be incorporated into an immunoassay. A chelating group that binds lanthanide ions is attached to antibody 2 in Figure 19-13. While bound to the antibody, Eu3+ has weak luminescence. After all steps in Figure 19-13 have been completed, the pH of the solution is lowered in the presence of a soluble chelator that extracts Eu3+ into solution. Strong luminescence from the soluble metal ion is then easily detected by a time-resolved measurement.

The idea of the use of luminescent lanthanide chelates in bioanalysis can be traced back to the 1980s, when Hemmila (1985) reported a time-resolved fluorometric system using an Eu labeling reagent, which was commercially produced by Wallac Oy company. The system was intended for immunoassay and included the spectrometer for time-resolved measurements. The basic principle of the immunoassay system (DELFIA ) is shown in fig. 1. 

Time-resolved lanthanide luminescence was used for the first time for immunoassays. This type of analytical method most effectively removes the background fluorescence, because serum contains many undesired co-existing materials, which raises significantly the background level when the sample is irradiated. 

Figure 13.2 Principles of (a) time-resolved spectroscopy, (h) heterogeneous immunoassays, and (c) homogeneous immunoassays [1]. (Reproduced from J.C.G Bunzli and C. Piguet, Taking advantage of luminescent lanthanide ions, Chemical Society Reviews, 34, 1048—1077, 2005, by permission of The Royal Society of Chemistry.)...

Pettersson K, Katajamaki T, Irjala K, Leppanen V, Majamaa-Volti K, Laitinen P. Time-resolved fluorom-etry (TRF)-based immunoassay concept for rapid and quantitative determination of biochemical myocardial infarction markers from whole blood, serum and plasma. Luminescence 2000 15 1-9. 

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