Preparation and identification of monoclonal antibody against enoyl-CoA hydratase 1 / 南方医科大学学报

ABSTRACT

<p><b>OBJECTIVE</b>To prepare monoclonal antibodies (mAbs) against enoyl-CoA hydratase 1 (ECH1).</p><p><b>METHODS</b>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.</p><p><b>RESULTS</b>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.</p><p><b>CONCLUSION</b>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.</p>