Dissociation-enhanced fluoroimmunoassay DELFIA

Fluoroimmunoassay makes use of the above behaviour. One of the common commercial methods is dissociation-enhanced fluoroimmunoassay (DELFIA). In this, a nonfluorescent Eu(III) EDTA-like complex is attached by a simple chemical reaction to an antibody or antigen, in a process called labelling. An immunoreaction is next initiated to bind the target, and then a (3-diketone and trioctylphosphine oxide (TOPO) mixture are added to the immunocomplex formed, at pH 3, to promote release of the Eu(III) from the antibody and its complexation as the strongly fluorescent complex [Eu((3-diketonate)3(TOPO)2], which is then measured by time-resolved fluorescence methods. The signal size relates to the amount of europium complexed, which in turns relates directly to the amount of the specifically formed target immunocomplex. This process is represented schematically in Figure 9.5. 

Fig. 25. (A) DELFIA (Dissociation Enhanced Lanthanide Fluoro-ImmunoAssay) system. This heterogeneous immunoassay system uses a primary antibody bound to a solid support, to which a variable amount of unlabeled antigen is bound. The secondary antibody is labeled with a non-phospho-rescent lanthanide chelate, which becomes phosphorescent after dissociation from the antibody, due to the addition of an enhancement solution [which typically contains a mixture of sensitizer (typically a (1-diketonate) and micelle inducing surfactant (5). (B) Heterogeneous fluoroimmunoassay using a secondary antibody directly labeled with a phosphorescent lanthanide chelate.

Schoket B, Doty WA, Vincze I, et al. 1993. Increased sensitivity for determination of polycyclic aromatic hydrocarbon-DNA adducts in human DNA samples by dissociation-enhanced lanthanide fluoroimmunoassay (DELFIA). Cancer Epidemiology, Biomarkers Prevention 2 349- 353. 

Several different assay formats, as already mentioned earlier, can generally be used such as ELISA, fluoroimmunoassay (FIA), dissociation-enhanced lanthanide fluoroimmunoassay (DELFIA), RIA, and SPR. For the quantification of proteins in biological matrices such as blood, plasma, serum or urine, most often FLISAs are validated. 

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