Ultrasensitive Time-Resolved Fluoroimmunoassay for Saikosaponin a in Chaihu (Bupleuri Radix).
Ultrasensitive Time-Resolved Fluoroimmunoassay for Saikosaponin a in Chaihu (Bupleuri Radix).
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超灵敏时间分辨荧光免疫分析柴胡中柴胡皂苷 a
DOI：
10.1371/journal.pone.0151032
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发表时间：
2016
期刊：
影响因子：
3.7
通讯作者：
Wu Y
中科院分区：
文献类型：
--
作者：
Chao Z;Cui Q;Tian E;Zeng W;Cai X;Li X;Tanaka H;Shoyama Y;Wu Y
建立柴胡药材中柴胡皂苷a （SSa）的时间分辨荧光免疫分析法（TRFIA）。用柴胡样品甲醇提取物和小鼠抗ssa单克隆抗体包被兔抗小鼠IgG的96孔微孔板孵育，以Eu3+标记的ssa -人血清白蛋白偶联物为示色剂。建立的竞争性TRFIA在0.01 ~ 10.0 μg/mL范围内具有良好的四阶多项式拟合，检测限为0.006 μg/mL。该方法的组内变异系数和组间变异系数分别为7.3%和8.9%，平均SSa回收率为119.2%。对于柴胡提取物样品，该方法的结果与之前建立的酶联免疫吸附法的结果具有良好的相关性。该系统对SSa的检测灵敏度高，检测范围宽，稳定性好，首次尝试用TRFIA对柴胡药材进行质量评价。
The aim of this study is to establish a time-resolved fluoroimmunoassay (TRFIA) system for quantitative analysis of saikosaponin a (SSa) in the crude drug of Chaihu (Bupleuri Radix). A 96-well microplate coated with rabbit anti-mouse IgG was incubated with the methanol extracts of Chaihu samples and a mouse anti-SSa monoclonal antibody, and a Eu3+-labeled SSa-human serum albumin conjugate was used as the tracer. The established competitive TRFIA showed a good fourth order polynomial fitting from 0.01 to 10.0 μg/mL for standard SSa sample with a detection limit of 0.006 μg/mL. The intra- and inter-assay coefficients of variation of the assay were 7.3% and 8.9%, respectively, and the average SSa recovery was 119.2%. For samples of Chaihu extract, the results of this assay showed a good correlation with those by enzyme-linked immunosorbent assay established previously. This TRFIA system is ultrasensitive for detecting SSa with a wide detection range and a good stability and represents the first attempt of using TRFIA for quality evaluation of the crude drug of Chaihu.