Vitamin D status can be determined by a radioimmunoassay measurement of 25-hydroxyvitamins D2 and D3 in plasma. For this purpose, 5 vitamin D haptens with different terminal carboxylic acid side chains were synthesised from vitamin D2. Rabbits, rats and mouse antibodies were generated by immunisation with bovine serum albumin conjugates of these 5 haptens. Histamine conjugates gave precursors of the radioiodinated tracers. 25-hydroxyvitamin D2 and D3 antibodies were obtained from all these animals with more or less equal recognition. Spleen cells from 3 rats immunised were fused with mouse myeloma cells. Two different hybridoma clones secreting monoclonal antibodies with equal response for the two vitamins and no cross-reaction with 1α,25-dihydroxyvitamin D were selected.
