INH表位多肽疫苗抗体间接ELISA测定方法的建立及优化

doi:10.16431/j.cnki.1671-7236.2016.08.031

《中国畜牧兽医》 ›› 2016, Vol. 43 ›› Issue (8): 2156-2163.doi: 10.16431/j.cnki.1671-7236.2016.08.031

INH表位多肽疫苗抗体间接ELISA测定方法的建立及优化

张玲玲¹, 岳耀敬¹, 冯瑞林¹, 李红峰², 郭婷婷¹, 袁超¹, 牛春娥¹, 刘建斌¹, 孙晓萍¹, 韩吉龙¹, 刘善博¹, 杨博辉¹

1. 中国农业科学院兰州畜牧与兽药研究所, 中国农业科学院羊育种工程技术研究中心, 兰州 730050;
2. 甘肃省武威市天祝县畜牧技术推广站, 天祝 733200

收稿日期:2015-11-23 出版日期:2016-08-20 发布日期:2016-08-23
通讯作者: 杨博辉 E-mail:yangbh2004@163.com
作者简介:张玲玲(1989-),女,河南开封人,硕士生,研究方向:动物繁殖调控,E-mail:15890388385@163.com
基金资助:
甘肃省青年自然基金“非繁殖季节GnIH基因免疫对藏羊卵泡发育的调节机制”；国家绒毛用羊产业技术体系分子育种岗位（CARS-40-03）；中国农业科学院“细毛羊资源与育种”创新工程（CAAS-ASTIP-2015-LIHPS）

Establishment and Optimization of INH Epitope Peptide Vaccine for Detection of Antibody

ZHANG Ling-ling¹, YUE Yao-jing¹, FENG Rui-lin¹, LI Hong-feng², GUO Ting-ting¹, YUAN Chao¹, NIU Chun-e¹, LIU Jian-bin¹, SUN Xiao-ping¹, HAN Ji-long¹, LIU Shan-bo¹, YANG Bo-hui¹

1. Lanzhou Institute of Animal Science & Veterinary Pharmaceutics, Chinese Academy of Agricultural Sciences, Lanzhou 730050, China;
2. Tianzhu County Animal Husbandry Technology Extending Stations, Tianzhu 733200, China

Received:2015-11-23 Online:2016-08-20 Published:2016-08-23

摘要/Abstract

摘要：

本试验在间接ELISA的基础上建立了一种检测抑制素（inhibit hormone, INH）表位多肽疫苗抗体的方法并对ELISA试剂盒进行优化，为今后测定主动免疫后细毛羊体内INH表位多肽疫苗抗体提供理论参考。在前人研究的基础上，采用间接ELISA法测定血清中INH表位多肽疫苗抗体的含量，通过对不同试验条件的控制摸索出最佳试验条件，即以脱脂奶粉为封闭液，INH及GnIH以表位多肽疫苗抗体稀释度为20 000倍，最佳反应时间60 min，最佳显色时间为15 min。本试验建立了一种检测细毛羊体内INH及GnIH抗体的方法，为今后对INH表位多肽疫苗抗体的研究提供了参考。

关键词: 抑制素; 细毛羊; 主动免疫; 生殖激素

Abstract:

In this study, an indirect ELISA method was established to detect inhibin hormone (INH) epitope peptide vaccine antibody, it would provide oretical reference for the determination of Fine-wool sheep after active immune body INH epitope peptide vaccine antibody. On the basis of the predecessors, using indirect ELISA method to determinate serum INH epitope peptide antibody levels of sheep, and through control different experimental conditions to look for the best experimental conditions. Through explorating the experimental conditions, finally, the testing experiment conditions were determined, which was blocked solution with skimmed milk powder, INH and GnIH synthetic peptides dilution degrees for 20 000 times, the optimum reaction time was 60 min, the best color action time was 15 min. In this experiment, a kind of method to detection antibody in the body after INH active immune sheep was built, it would provide a reference for future research.

Key words: inhibin hormone; Fine-wool sheep; active immunization; reproductive hormone

中图分类号:

S852.4

张玲玲, 岳耀敬, 冯瑞林, 李红峰, 郭婷婷, 袁超, 牛春娥, 刘建斌, 孙晓萍, 韩吉龙, 刘善博, 杨博辉. INH表位多肽疫苗抗体间接ELISA测定方法的建立及优化[J]. 《中国畜牧兽医》, 2016, 43(8): 2156-2163.

ZHANG Ling-ling, YUE Yao-jing, FENG Rui-lin, LI Hong-feng, GUO Ting-ting, YUAN Chao, NIU Chun-e, LIU Jian-bin, SUN Xiao-ping, HAN Ji-long, LIU Shan-bo, YANG Bo-hui. Establishment and Optimization of INH Epitope Peptide Vaccine for Detection of Antibody[J]. , 2016, 43(8): 2156-2163.

参考文献

[1] 茆达干,杨利国,姜勋平,等.抑制素α(1~32)基因免疫对大鼠卵泡发育和生殖激素的影响[J].中国农业科学,2003,36(12):1154-1159.
[2] 茆达干,杨利国,张志杰,等.抑制素基因免疫的免疫反应性[J].中国兽医学报,2006,26(3):292-295.
[3] 姜勋平,杨利国,刘桂琼,等.抑制素基因免疫对小鼠生殖的影响[J].中国兽医学报,2002,22(4):368-369.
[4] Cummmins J M.Seprm size body massandre production inmammals[J].Seprmcell,1983,114(4):395-398.
[5] Findlay J K,Clarke I J,Robertson D M.Inhibin concentrations in ovarian and jugularvenous plasmaand the relationship of inhibin with follicle-stimulating hormone and luteinizing hormone during the ovineestrous cycle[J].Endocrinology,1990,126(1):528-535.
[6] Tsutsui K.Anewkey neuro hormone control ling reproduction,gonadotrop-ininhibit oryhormone (GnIH):Biosynthesis,mode of action and functional significance[J].Progressin Neurobiology,2009,88(1):76-88.
[7] Bartlewski P M,Beard A P,Cook S J,et al.Ovarian antral follicular dynamics and their relationships with endocrine evariables throughout the estrous cycle in breeds of sheep differing in prolificacy[J].J Reprod Fertil,2011,115(1):111-124.
[8] Clarke I J,Qi Y,Puspita S I,et al.Evidence that RF-amide related peptide sare inhibitors of reproduction in mammals[J].Frontiersin Neuroendocrinology, 2009,30(3):371-378.
[9] Condon T P,Leipheimer R E,Curry J J.Preliminary evidence for a CNS site of action for ovarian inhibin[J].Life Sciences, 2010,32(15):1691-1698.
[10] Miller W L,Huang E S.Secretion of ovine luteinzing hormone in vitro:Differential positive control by 17-β estraiol and a preparation of porcine ovarian inhibin[J].Endocr,1985,117(3):907-901.
[11] Wrathall J E.Inhibin immunoneut ralization by antibodis raised against synthetic peptide sequences of inhibin α-subunit effect son gonadotrophin concentrations and ovulation rate in sheep[J].Endocrinal,1990,124:167-176.
[12] Leizhihat L Y.Recent studies of gonadotropin-inhibitory hormone[J].Animal Husbandry & Veterinary Medicine,2008,40(9):60-100.
[13] 李万宏,王春强,周虚.抑制素对动物生殖系统的影响及其在动物生产中的应用[J].中国畜牧兽医,2011,38(6):204-207.
[14] Ding H Z,Yu B S.Relationship between gonadotropin-inhibitory hormone and hypothalamus-pituitary-gonadalaxis[J].Progressin ModernBiomedicine,2010,10(23):4598-4600.
[15] Hinuma S,Shintani Y,Fukusumi S,et al.New neuro peptides containing carboxy-terminal RF-amide and the irreceptor in mammals[J].Nat Cell Biol,2000,2(10):703-708.
[16] Ukena K,Iwakoshi E,Minakata H,et al.A novel rat hypothalamic RFamide-related peptide identified by immune affinity chromate graphy and mass spectrometry[J].FEBS Lett,2002,512(1-3):255-258.
[17] 焦彩兰,储明星,王金玉.绵羊高繁殖力内分泌学机制的研究进展[J].中国畜牧兽医,2006,33(1):30-32.
[18] Hennies M,Voglmayr J K,Dietrich E,et al.Hormonal resonse of femal goat stoactive immunization against are combinant human inhibin subunit,and establishment of an enzyme linked immune sorbernt assay for caprine follicle stimulating hormone[J].Reprod Dom Anim,2001,36(2):65-71.
[19] Naqvi S M,Gulyani R.The effect of gonadotrophin releasing hormone and follicle stimulating hormone in conjunction with pregnant mareserum gonadotrophinon the super ovulatory response in crossbred sheep in India[J].Trop Anim Health Prod,1998,30(6):369-376.
[20] 王水莲,薛立群,邢朝芳,等.卵泡抑制素与GFP融合基因疫苗免疫对大鼠卵巢和生殖激素的影响[J].中国兽医学报,2012,32(2):309-313.
[21] 王水莲,冯建忠,李文平,等.非抗性筛选抑制素基因疫苗免疫免疫对奶牛产后子宫复旧的影响[J].湖南大学农业学报,2014,40(4):395-398.

INH表位多肽疫苗抗体间接ELISA测定方法的建立及优化

Establishment and Optimization of INH Epitope Peptide Vaccine for Detection of Antibody

PDF

可视化

摘要/Abstract

引用本文

使用本文

参考文献

相关文章 15

编辑推荐

Metrics

本文评价

[1]	马继福, 阿德力·克坦, 解津刚, 巴合提·博代, 李振伟, 吾热力哈孜·哈孜汗. 基于噬菌体展示技术筛选抑制素α亚基特异性纳米抗体[J]. 中国畜牧兽医, 2023, 50(5): 2032-2043.
[2]	马继福, 巴合提·博代, 黑子别克·哈孜别克, 李振伟, 吾热力哈孜·哈孜汗. 抑制素α亚基特异性纳米抗体基因筛选与鉴定[J]. 中国畜牧兽医, 2023, 50(4): 1532-1542.
[3]	王志亮, 杨喜喜, 铁雅楠, 伏晓玉, 王欣荣. 超排处理对家兔生殖器官形态及激素水平的影响[J]. 中国畜牧兽医, 2022, 49(6): 2250-2258.
[4]	梁家充, 吕春荣, 洪琼花, 吴国权, 权国波. 云南半细毛羊精子冷冻前后转录组表达差异分析[J]. 中国畜牧兽医, 2022, 49(3): 973-981.
[5]	赫雪, 谷英, 斯登丹巴, 德德玛, 窦骁宁, 吴怡, 雷志惠, 潘和平, 岳耀敬. 基于单细胞转录组测序技术对鄂尔多斯细毛羊次级毛囊形态发生诱导期的研究[J]. 中国畜牧兽医, 2022, 49(11): 4296-4306.
[6]	李芝丰, 储明星, 孙伟. 绵羊INHBB、SMAD4和FGF18基因的双荧光素酶载体构建及其与miR-370-3p的靶向验证[J]. 中国畜牧兽医, 2021, 48(9): 3109-3117.
[7]	苗曼宁, 郭益文, 胡德宝, 曾雨晗, 李新, 张林林, 丁向彬, 郭宏. MSTN基因编辑牛肌肉转录组测序及生物信息学分析[J]. 中国畜牧兽医, 2021, 48(7): 2271-2281.
[8]	贾银海, 黄明光, 徐文文, 李芳芳, 文信旺, 李铭, 杨秀荣, 吴柱月, 蒋和生. 水牛发情期唾液生殖激素和结晶与卵泡发育的关系[J]. 中国畜牧兽医, 2021, 48(2): 592-598.
[9]	丽娜·沙肯, 巴合提·博代, 杨华, 解津刚, 董巧燕, 吾热力哈孜·哈孜汗. 抑制素α主动和被动免疫对哈萨克羊生殖激素含量的影响[J]. 中国畜牧兽医, 2021, 48(2): 658-667.
[10]	盛辉, 郭益文, 张林林, 谭皓云, 胡德宝, 李新, 丁向彬, 郭宏. MSTN基因对牛肌动蛋白细胞骨架调节通路的影响[J]. 中国畜牧兽医, 2021, 48(10): 3554-3564.
[11]	范玉珊, 吴俊辉, 高凤磊, 郭聪慧, 谢绍怡, 孟立, 李莉, 张守全, 卫恒习. 定时输精对经产母猪繁殖性能及生殖激素的影响[J]. 中国畜牧兽医, 2021, 48(10): 3726-3733.
[12]	王涛, 李玉荣, 何云梅, 胥福勋, 孙雪峰, 阿拉木斯, 孙春河, 赵生国, 刘斌. 诱导发情对绒山羊非繁殖季节产羔和血清指标的影响[J]. 中国畜牧兽医, 2020, 47(6): 1819-1827.
[13]	盛辉, 张林林, 郭益文, 李新, 丁向彬, 郭宏. 干扰MSTN的牛骨骼肌卫星细胞实时荧光定量PCR内参基因的筛选[J]. 中国畜牧兽医, 2020, 47(2): 381-391.
[14]	欧阳依娜, 李银江, 李卫娟, 薛白, 王思宇, 梁家充, 洪琼花. 不同钙水平日粮对云南半细毛羊泌乳期能量代谢的影响[J]. 中国畜牧兽医, 2020, 47(2): 416-424.
[15]	朱菲菲, 张俊星, 张林林, 李新, 刘新峰, 郭益文, 丁向彬. 干扰MSTN对牛骨骼肌卫星细胞增殖分化的影响[J]. 中国畜牧兽医, 2020, 47(2): 479-487.