Production and characterization of monoclonal antibodies against a novel 1alpha,25-dihydroxyvitamin D3-bovine serum albumin conjugate linked through the 11alpha-position

The Journal of Steroid Biochemistry and Molecular Biology
Norihiro KobayashiK Shimada

Abstract

To obtain an antibody which is useful for developing a selective immunoaffinity chromatography (IAC) of 1alpha,25-dihydroxyvitamin D3 [1,25(OH)2D3], which is an effective pretreatment tool for determining this metabolite in biological fluids, a variety of monoclonal antibodies have been produced against a novel haptenic derivative, 11alpha-hemiglutaryloxy-1alpha,25-dihydroxyvitamin D3, linked to bovine serum albumin. BALB/c, A/J and C57BL/6 mice, as well as Wistar and SD rats, were immunized with the hapten-carrier conjugate and the titer of the anti-1,25(OH)2D3 antibody in serum was examined. Spleen cells from A/J mice or SD rats, which showed a high titer, were fused with P3/NS1/1-Ag4-1 myeloma cells. After serial screening by enzyme-linked immunosorbent assay and radioimmunoassay, sixteen kinds of hybridoma clones secreting different anti-1,25(OH)2D3 antibodies were established. Scatchard analysis revealed that 12 antibodies out of 16 have enough affinity to 1,25(OH)2D3 (Ka>10(9) M[-1]). By a cross-reaction study using various related compounds, two antibodies, M114 and M129, both derived from A/J mice, were found to have suitable specificity. Although they showed relatively high cross-reactivities with the 1,25(OH)2D3 deriv...Continue Reading

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Citations

Dec 28, 2002·Analytical Sciences : the International Journal of the Japan Society for Analytical Chemistry·Norihiro KobayashiHiroshi Hosoda
Jan 10, 2018·Biological & Pharmaceutical Bulletin·Izumi MoritaNorihiro Kobayashi
Feb 15, 2003·The Journal of Organic Chemistry·Lars K A BlaehrMikael Begtrup

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