Department of Microbiology, Faculty of Biotechnology and Biomolecular Sciences, Universiti Putra Malaysia, 43400 Serdang, Selangor, Malaysia.
J Chromatogr A. 2010 May 21;1217(21):3473-80. doi: 10.1016/j.chroma.2010.03.012. Epub 2010 Mar 18.
Hepatitis B core antigen (HBcAg) is used as a diagnostic reagent for the detection of hepatitis B virus infection. In this study, immobilized metal affinity-expanded bed adsorption chromatography (IMA-EBAC) was employed to purify N-terminally His-tagged HBcAg from unclarified bacterial homogenate. Streamline Chelating was used as the adsorbent and the batch adsorption experiment showed that the optimal binding pH of His-tagged HBcAg was 8.0 with a binding capacity of 1.8 mg per ml of adsorbent. The optimal elution condition for the elution of His-tagged HBcAg from the adsorbent was at pH 7 in the presence of 500 mM imidazole and 1.5 M NaCl. The IMA-EBAC has successfully recovered 56% of His-tagged HBcAg from the unclarified E. coli homogenate with a purification factor of 3.64. Enzyme-linked immunosorbent assay (ELISA) showed that the antigenicity of the recovered His-tagged HBcAg was not affected throughout the IMA-EBAC purification process and electron microscopy revealed that the protein assembled into virus-like particles (VLP).
乙型肝炎核心抗原（HBcAg）被用作检测乙型肝炎病毒感染的诊断试剂。在本研究中，固定化金属亲和扩展床吸附色谱（IMA-EBAC）被用于从未澄清的细菌匀浆中纯化 N 端组氨酸标签的 HBcAg。Streamline Chelating 被用作吸附剂，批量吸附实验表明，His 标记的 HBcAg 的最佳结合 pH 值为 8.0，结合容量为 1.8 mg/ml 吸附剂。从吸附剂上洗脱 His 标记的 HBcAg 的最佳洗脱条件为 pH 7，存在 500 mM 咪唑和 1.5 M NaCl。IMA-EBAC 已成功从未澄清的大肠杆菌匀浆中回收了 56%的 His 标记的 HBcAg，纯化倍数为 3.64。酶联免疫吸附试验（ELISA）表明，回收的 His 标记的 HBcAg 的抗原性在整个 IMA-EBAC 纯化过程中不受影响，电子显微镜显示该蛋白组装成病毒样颗粒（VLP）。
