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Enzyme-linked immunosorbent assay of changes in serum levels of growth hormone (cGH) in common carps (Cyprinus carpio)
Wu, Gang1; Chen, LiHua2; Zhong, Shan1,3; Li, Qi2; Song, ChaoJun2; Jin, BoQuan2; Zhu, ZuoYan1; Zhu, ZY, Wuhan Univ, Chinese Acad Sci, Inst Hydrobiol, State Key Lab Freshwater Ecol & Biotechnol, Wuhan 430072, Peoples R China
2008-02-01
Source PublicationSCIENCE IN CHINA SERIES C-LIFE SCIENCES
ISSN1006-9305
Volume51Issue:2Pages:157-163
AbstractThe aim of the present study was to purify the common native carp growth hormone (ncGH), produce monoclonal antibodies (mAbs) to common native carp growth hormone (ncGH), and further enhance the sensitivity of enzyme-linked immunosorbent assays (ELISA) for ncGH. Additionally, we investigated changes in serum ncGH levels in carps raised in different environmental conditions. The recombinant grass carp (Ctenopharyngodon idella) growth hormone was purified and used as antigen to immunize the rabbit. The natural ncGH was isolated from the pituitaries of common carp. SDS-PAGE and Western blot utilizing the polyclonal anti-rgcGH antibody confirmed the purification of ncGH from pituitaries. Purified ncGH was then used as an immunogen in the B lymphocyte hybridoma technique. A total of 14 hybridoma cell lines (FMU-cGH 1-14) were established that were able to stably secrete mAbs against ncGH. Among them, eight clones (FMU-cGH1-6, 12 and 13) were successfully used for Western blot while nine clones (FMU-cGH 1-7, 9 and 10) were used in fluorescent staining and immunohistochemistry. Epitope mapping by competitive ELISA demonstrated that these mAbs recognized five different epitopes. A sensitive sandwich ELISA for detection of ncGH was developed using FMU-cGH12 as the coating mAb and FMU-cGH6 as the enzyme labeled mAb. This detection system was found to be highly stable and sensitive, with detection levels of 70 pg/mL. Additionally, we found that serum ncGH levels in restricted food group and in the net cage group increased 6.9-and 5.8-fold, respectively, when compared to controls, demonstrating differences in the GH stress response in common carp under different living conditions.; The aim of the present study was to purify the common native carp growth hormone (ncGH), produce monoclonal antibodies (mAbs) to common native carp growth hormone (ncGH), and further enhance the sensitivity of enzyme-linked immunosorbent assays (ELISA) for ncGH. Additionally, we investigated changes in serum ncGH levels in carps raised in different environmental conditions. The recombinant grass carp (Ctenopharyngodon idella) growth hormone was purified and used as antigen to immunize the rabbit. The natural ncGH was isolated from the pituitaries of common carp. SDS-PAGE and Western blot utilizing the polyclonal anti-rgcGH antibody confirmed the purification of ncGH from pituitaries. Purified ncGH was then used as an immunogen in the B lymphocyte hybridoma technique. A total of 14 hybridoma cell lines (FMU-cGH 1-14) were established that were able to stably secrete mAbs against ncGH. Among them, eight clones (FMU-cGH1-6, 12 and 13) were successfully used for Western blot while nine clones (FMU-cGH 1-7, 9 and 10) were used in fluorescent staining and immunohistochemistry. Epitope mapping by competitive ELISA demonstrated that these mAbs recognized five different epitopes. A sensitive sandwich ELISA for detection of ncGH was developed using FMU-cGH12 as the coating mAb and FMU-cGH6 as the enzyme labeled mAb. This detection system was found to be highly stable and sensitive, with detection levels of 70 pg/mL. Additionally, we found that serum ncGH levels in restricted food group and in the net cage group increased 6.9-and 5.8-fold, respectively, when compared to controls, demonstrating differences in the GH stress response in common carp under different living conditions.
SubtypeArticle
KeywordCommon Carp Growth Hormone Monoclonal Antibodies Elisa Starvation Net Cage
Department[Wu, Gang; Zhong, Shan; Zhu, ZuoYan] Wuhan Univ, Chinese Acad Sci, Inst Hydrobiol, State Key Lab Freshwater Ecol & Biotechnol, Wuhan 430072, Peoples R China; [Chen, LiHua; Li, Qi; Song, ChaoJun; Jin, BoQuan] Fourth Mil Med Univ, Dept Immunol, Xian 710032, Peoples R China; [Zhong, Shan] Wuhan Univ, Coll Life Sci, Wuhan 430072, Peoples R China
Subject AreaBiology
DOI10.1007/s11427-008-0022-z
WOS HeadingsScience & Technology ; Life Sciences & Biomedicine
Indexed BySCI
Language英语
WOS Research AreaLife Sciences & Biomedicine - Other Topics
WOS SubjectBiology
WOS IDWOS:000252876000009
WOS KeywordCONFINEMENT STRESS ; RADIOIMMUNOASSAY ; VALIDATION ; RECEPTOR ; FISH ; PITUITARY ; LIVER
Citation statistics
Cited Times:12[WOS]   [WOS Record]     [Related Records in WOS]
Document Type期刊论文
Identifierhttp://ir.ihb.ac.cn/handle/152342/8284
Collection期刊论文
Corresponding AuthorZhu, ZY, Wuhan Univ, Chinese Acad Sci, Inst Hydrobiol, State Key Lab Freshwater Ecol & Biotechnol, Wuhan 430072, Peoples R China
Affiliation1.Wuhan Univ, Chinese Acad Sci, Inst Hydrobiol, State Key Lab Freshwater Ecol & Biotechnol, Wuhan 430072, Peoples R China
2.Fourth Mil Med Univ, Dept Immunol, Xian 710032, Peoples R China
3.Wuhan Univ, Coll Life Sci, Wuhan 430072, Peoples R China
Recommended Citation
GB/T 7714
Wu, Gang,Chen, LiHua,Zhong, Shan,et al. Enzyme-linked immunosorbent assay of changes in serum levels of growth hormone (cGH) in common carps (Cyprinus carpio)[J]. SCIENCE IN CHINA SERIES C-LIFE SCIENCES,2008,51(2):157-163.
APA Wu, Gang.,Chen, LiHua.,Zhong, Shan.,Li, Qi.,Song, ChaoJun.,...&Zhu, ZY, Wuhan Univ, Chinese Acad Sci, Inst Hydrobiol, State Key Lab Freshwater Ecol & Biotechnol, Wuhan 430072, Peoples R China.(2008).Enzyme-linked immunosorbent assay of changes in serum levels of growth hormone (cGH) in common carps (Cyprinus carpio).SCIENCE IN CHINA SERIES C-LIFE SCIENCES,51(2),157-163.
MLA Wu, Gang,et al."Enzyme-linked immunosorbent assay of changes in serum levels of growth hormone (cGH) in common carps (Cyprinus carpio)".SCIENCE IN CHINA SERIES C-LIFE SCIENCES 51.2(2008):157-163.
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