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Enzyme-linked immunosorbent assay of changes in serum levels of growth hormone(cGH)in common carps(Cyprinus carpio) 被引量:1

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摘要 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. 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.
出处 《Science China(Life Sciences)》 SCIE CAS 2008年第2期157-163,共7页 中国科学(生命科学英文版)
基金 the Development Plan of the State Key Fundamental Research ofChina(Grant No.2007CB109206) Chinese National Programs for High Technology Research and Development(Grant No.2006AA10Z141) the National NaturalScience Foundation of China(Grant No.30130050)
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  • 1Sumpter J P,Le Bail P Y,Pickering A D,et al.The effect of starvation on growth and plasma growth hormone concentration of rainbow trout,Oncorhynchus mykiss. General and Comparative Endocrinology . 1991
  • 2Deng L,Zhang W M,Lin H R,et al.Effects of food deprivation on expression of growth hormone receptor and proximate composition in liver of black seabream Acanthopagrus schlegeli. Comparative Biochemistry and Physiology . 2004
  • 3Carter-Su C,Schwartz J,Smit L S.Molecular mechanism of growth hormone action. Annu Rev Phys . 1996
  • 4Reinecke M,Bjornsson B T,Dickhoff WW,et al.Growth hormone and insulin-like growth factors in fish:Where we are and where to go. General and Comparative Endocrinology . 2005
  • 5Waters M J,Rowlinson S W,Clarkson R W,et al.Signal transduction by the growth hormone receptor. Proceedings of the Society for Experimental Biology and Medicine . 1994
  • 6Mertani H C,Waters M J,Jambou R,et al.Growth hormone receptor binding protein in rat anterior pituitary. Neuroendocrinology . 1994
  • 7Rotllant J,Balm P H,Perez-Sanchez J,et al.Pituitary and interrenal function in gilthead sea bream(Sparus aurata L.Teleostei)after handling and confinement stress. General and Comparative Endocrinology . 2001
  • 8Wabitsch M,Hauner H,Heinze E,Teller WM.The role of growth hormone/insulin- like growth factor in adipocyte differentiation. Metabolism . 1995
  • 9Perez Sanchez J,Smal J,Le Bail P Y.Location and characterization of growth hormone binding sites in the central nervous system of a teleost fish(Oncorhynchus mykiss). Growth Regulation . 1992
  • 10Sun,X,Zhu,S,Chan,SSH,Toresson,G,Cheng,CHK.Identification and characterization of growth hormone receptors in snakehead fish (Ophiocephalus argus Cantor) liver. General and Comparative Endocrinology . 1997

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