ELISA for the Determination of Saikosaponin a, an Active Component of Bupleuri Radix

概要

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In order to quantify saikosaponin a (SSA), one of the major active components of Bupleuri Radix, a competitive and indirect ELISA method was developed.High titer rabbit polyclonal antibodies (pAbs) were raised against a conjugate of SSA and bovine serum albumin, coupled with a periodate oxidation method. SSA competitively inhibited the binding of rabbit anti-SSA pAbs to SSA-ovalbumin on the solid phase, a coated antigen on the well. The quantity of pAbs bound to the well was monitored using a peroxidase-conjugated anti-rabbit IgG as a secondary antibody, and tetramethylbenzidine solution as a substrate. The measuring range extended from 50 pg/ml to 20 ng/ml of SSA, with a detection limit of 40 pg/ml (5.13 pM). Antibodies showed some cross-reactivity with saikosaponin c (12.74%). However, the antibodies showed only slight cross-reactivities with saikosaponin d (0.3%), which differs from SSA only in the stereochemistry of the 16-hydroxyl group, and the artificial saikosaponins, saikosaponin b_1 (2.1%) and saikosaponin g (0.53%). The specific and sensitive ELISA is especially suited for determination of SSA in samples when only small quantities of materials can be extracted for analysis.
公益社団法人日本薬学会の論文
1998-07-15