The antibody is an affinity purified rabbit polyclonal IgG (supplied by New England Biolabs). Affinities for the various phosphothreonine peptides are as shown below. Affinities are expressed as signal-to-noise ratios measured using 10ng/well anti-phosphothreonine antibody followed by DELFIA® europium labelled anti-rabbit antibody for detection. Noise is the reactivity with the non-phosphopeptide.

Generic DELFIA® reagents are intended for use in dissociation-enhanced time-resolved fluorometric assays. All reagents have been labeled using the DELFIA® N1-chelate. They are suitable for highly sensitive end point measurements in separation based assays such as those for detection of protein-protein binding and cell adhesion as well as for immunoassays.

DELFIA® (dissociation-enhanced lanthanide fluorescence immunoassay) is a time-resolved fluorescence (TRF) intensity technology. Assays are designed to detect the presence of a compound or biomolecule using lanthanide chelate labeled reagents, separating unbound reagent using wash steps. DELFIA assays are flexible, compatible with a variety of plate readers, and, as this is a wash-based technology, compatible with most sample types. The technology is based on fluorescence of lanthanide chelates (Europium, Samarium, and Terbium). The fluorescence decay time of these lanthanide chelate labels is much longer than traditional fluorophores, allowing efficient use of temporal resolution for reduction of autofluorescent background. The large Stokes’ shift (difference between excitation and emission wavelengths) and the narrow emission peaks contribute to increasing signal-to-noise ratio. Sensitivity is further increased because of the dissociation-enhancement principle: the lanthanide chelate is dissociated and a new highly fluorescent chelate is formed into a protective micellar solution. DELFIA lanthanide chelates require this dissociation/enhancement step for fluorescence (induced by addition of DELFIA Enhancement solution, DELFIA Inducer, and DELFIA Enhancer as appropriate to the particular lanthanide chelate).