[Generation and characterization of monoclonal antibody against HSD11B1].
OBJECTIVE
To generate and identify monoclonal antibodies (mAb) against homo sapiens hydroxysteroid 11-beta dehydrogenase 1 (HSD11B1).


METHODS
Normal human liver tissues were homogenized, and mitochondria were isolated by differential centrifugation. The total mitochondrial proteins were used to immunize BALB/c mice to generate mAb by hybridoma technique. The mAb were characterized by ELISA, Western blot and immunohistochemistry. The antibody specificity was identified by immunoprecipitation (IP), and confirmed by Uni-ZAP expression library screening.


RESULTS
One hybridoma CBF245 secreting specific mAb against HSD11B1 was established. The Ig subclass of this mAb was IgG1, and it could be used in ELISA, Western blot, immunohistochemistry. Our data showed that the antigen recognized by CBF245 mAb was localized in the hepatocyte cytoplasm, with molecular weight of M(r) 35 000 in the mitochondria of human liver tissue. The CBF245 mAb was further confirmed by immunoscreening of Uni-ZAP XR liver cDNA expression library.


CONCLUSIONS
A hybridoma cell line stably secreting specific mAb against HSD11B1 is established and characterized. This mAb reacted with HSD11B1 in ELISA, Western blot, immunohistochemistry assay, IP, and would be very useful for the HSD11B1 studies.
Wang
Wan
W
Department of Immunology, Institute of Radiation Medicine, Beijing 100850, China. wangwanz@sina.com
Yang
Jin-ju
JJ
Liu
Li
L
Chen
Zhi-cheng
ZC
Gao
Yuan
Y
Gao
Jian-en
JE
An
Li-guo
LG
Sun
Qi-hong
QH
D043205:11-beta-Hydroxysteroid Dehydrogenase Type 1 / Q000276:immunology; D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis* / Q000276:immunology; D000918:Antibody Specificity; D006801:Humans; D006825:Hybridomas / Q000276:immunology*; D007075:Immunoglobulin M / Q000276:immunology; D007150:Immunohistochemistry; D051379:Mice
D000911:Antibodies, Monoclonal; D007075:Immunoglobulin M; D043205:11-beta-Hydroxysteroid Dehydrogenase Type 1; C468386:HSD11B1 protein, human