[Preparation and identification of monoclonal antibody against enoyl-CoA hydratase 1].
OBJECTIVE
To prepare monoclonal antibodies (mAbs) against enoyl-CoA hydratase 1 (ECH1).


METHODS
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.


RESULTS
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.


CONCLUSIONS
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.
Ju
Yan-fang
YF
General Hospital of PLA, Beijing 100853, China. yanfangju@yahoo.com.cn
Liu
Rong
R
Liu
Xiao-lan
XL
Yang
Jin-ju
JJ
Gao
Jian-en
JE
Sun
Qi-hong
QH
D000818:Animals; D000911:Antibodies, Monoclonal / Q000032:analysis* / Q000276:immunology*; D000918:Antibody Specificity; D015153:Blotting, Western; D002460:Cell Line; D004746:Enoyl-CoA Hydratase / Q000276:immunology*; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D007150:Immunohistochemistry; D008099:Liver / Q000166:cytology / Q000378:metabolism; D051379:Mice; D008807:Mice, Inbred BALB C; D008928:Mitochondria / Q000378:metabolism
D000911:Antibodies, Monoclonal; D004746:Enoyl-CoA Hydratase