Ju Yan-fang, Liu Rong, Liu Xiao-lan, Yang Jin-ju, Gao Jian-en, Sun Qi-hong
General Hospital of PLA, Beijing 100853, China.
Nan Fang Yi Ke Da Xue Xue Bao. 2009 Apr;29(4):648-51.
To prepare monoclonal antibodies (mAbs) against enoyl-CoA hydratase 1 (ECH1).
Normal human liver tissues were homogenized, and the mitochondria were isolated by differential centrifugation. The total mitochondrial proteins were used to immunize BALB/c mice to prepare mAbs by routine hybridoma technique. The mAbs were characterized by ELISA, Western blotting and immunohistochemistry. The specificity of the antibody was identified by mass spectrometry (MS) following immunoprecipitation (IP) and confirmed by Uni-ZAP expression library screening.
One clone of the hybridoma BGB095 secreting specific mAb against ECH1 was obtained. The mAb was identified to belong to Ig subclass IgG1 and could be used in ELISA, Western blotting, immunohistochemistry, and immunoprecipitation.
A hybridoma cell line stably secreting specific mAb against ECH1 has been established. The specific mAb against ECH1 can be of great value for functional and distribution studies of ECH1.
制备抗烯酰辅酶A水合酶1（ECH1）的单克隆抗体（mAb）。
将正常人肝组织匀浆，通过差速离心分离线粒体。用线粒体总蛋白免疫BALB/c小鼠，采用常规杂交瘤技术制备单克隆抗体。通过酶联免疫吸附测定（ELISA）、蛋白质免疫印迹法（Western blotting）和免疫组织化学法对单克隆抗体进行鉴定。在免疫沉淀（IP）后通过质谱（MS）鉴定抗体的特异性，并通过Uni-ZAP表达文库筛选加以证实。
获得了一株分泌抗ECH1特异性单克隆抗体的杂交瘤BGB095。该单克隆抗体被鉴定属于Ig亚类IgG1，可用于ELISA、Western blotting、免疫组织化学和免疫沉淀。
已建立稳定分泌抗ECH1特异性单克隆抗体的杂交瘤细胞系。抗ECH1特异性单克隆抗体对ECH1的功能和分布研究具有重要价值。
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