A One-Step Enzyme-Linked Immunosorbent Assay for a Novel Osteoblast Differentiation-Promoting Compound, TAK-778 in Serum

抄録

TAK-778, ((2R, 4S)-(-)-N-[(4-diethoxyphosphorylmethyl)phenyl]-1, 2, 4, 5-tetrahydro-4-methyl-7, 8-methylene-dioxy-5-oxo-3-benzothiepin-2-carboxamide), is a novel osteoblast differentiation-promoting compound, and its sustained-release formulation is expected to be clinically used for the enhancement of fracture healing. To date, TAK-778 levels in serum have been measured using conventional reverse-phase HPLC with inferior sensitivity and time-consuming procedures. We have produced polyclonal antibodies against TAK-778 by using one of its derivatives coupled with a carrier protein, and developed a one-step ELISA for the determination of TAK-778 in serum. The antibodies had minimal cross-reactivities to the biologically inactive metabolites including the oxidized-form (0.36%) and the cleavaged-form (0.00083%). The competitive ELISA was accomplished within three hours with a detection limit of 0.5ng/ml serum, and the coefficients of variations for samples ranging from 1 ng/ml to 80 ng/ml were 1.1-4.4% in an intra-assay and 3.1-9.6% in an inter-assay. The present ELISA is so rapid, sensitive and selective for TAK-778 that it could be conveniently used in a clinical field for the determination of many serum specimens.