A Monoclonal Antibody-Based Enzyme-Linked Immunosorbent Assay for Determination of Homoharringtonine
A Monoclonal Antibody-Based Enzyme-Linked Immunosorbent Assay for Determination of Homoharringtonine
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DOI：
10.1055/a-0578-8689
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发表时间：
2018-09-01
期刊：
影响因子：
2.7
通讯作者：
Morimoto, Satoshi
中科院分区：
文献类型：
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作者：
Pongkitwitoon, Benyakan;Sakamoto, Seiichi;Morimoto, Satoshi
高三尖杉酯碱（HHT），也称为奥马西他辛，是在三尖杉属中发现的天然化合物，是一种有前途的药物，用于治疗慢性或加速期慢性粒细胞白血病。作为定量测定 HHT 的工具，通过高碘酸钠介导的氧化制备的缀合物产生了针对 HHT 的特异性单克隆抗体 (MAb 6A1)。使用 MAb 6A1 开发的间接竞争性酶联免疫吸附测定 (icELISA) 具有高度特异性和敏感性，HHT 的检测限为 48.8ng/mL。通过使用测定内和测定间分析、回收率测试以及通过 ELISA 和高效液相色谱法测定的 HHT 量之间的比较分析，进行了验证测定，以评估该方法的精密度和准确度。这些结果表明，使用 MAb 6A1 建立的 icELISA 特异性、灵敏且可靠，足以应用于 HHT 的定性分析。此外，本研究的结果支持高碘酸钠作为三尖杉生物碱与蛋白质缀合以产生特异性抗体的试剂的有用性。
Homoharringtonine (HHT), also known as omacetaxine, is a natural compound found in the genus Cephalotaxus and is a promising pharmaceutical drug used for the treatment of chronic or accelerated phase chronic myeloid leukemia. As a tool for the quantitative determination of HHT, a specific monoclonal antibody against HHT (MAb 6A1) was generated by conjugates prepared via sodium periodate-mediated oxidation. The developed indirect competitive enzyme-linked immunosorbent assay (icELISA) using MAb 6A1 was found to be highly specific and sensitive with a limit of detection for HHT of 48.8ng/mL. Validation assays to evaluate precision and accuracy of the method were conducted by the use of intra- and inter-assay analysis, recovery test, and comparison analysis between the amounts of HHT determined by ELISA and high-performance liquid chromatography. These results revealed that the established icELISA using MAb 6A1 is specific, sensitive, and reliable enough to be applied to the qualitative analysis for HHT. Furthermore, the results of this study support the usefulness of sodium periodate as a reagent for the conjugation between Cephalotaxus alkaloids and proteins for producing specific antibodies.