HA蛋白多肽抗体的设计、制备与鉴定  

Design,Preparation and Identification of HA Peptide Antibody

作　　者：袁红花[1] 王珍珍[1] 陈仁金[1] 胡安康[1] 张腾业 吴连连[1] 冀德君 朱孝荣[1] 

机构地区：[1]徐州医学院,江苏徐州221004 [2]扬州大学,江苏扬州225009

出　　处：《家畜生态学报》2013年第12期18-21,共4页Journal of Domestic Animal Ecology

基　　金：国家自然科学基金(31172171);江苏省青年基金(BK2012138);江苏省自然科学基金(BK2011209);徐州医学院院长基金(2012KJZ20)

摘　　要：利用人工合成的多肽制备HA的特异性多克隆抗体,以期用于HA基因的免疫调控机制研究。根据GenBank中报道的皮蝇素A(Hydermin A,HA)一级结构信息以及HA抗原生物学信息,获得三段序列特异性较高的多肽,采用9一氟甲氧羰基固相合成法获得多肽,采用HPLC和LC·MS测定合成多肽的浓度和分子量,试验表明目的多肽纯度达91.2%、目的多肽分子量为28kD。将多肽与KLH进行偶联,并用其免疫新西兰大白兔以获得抗血清和多克隆抗体。采用ELISA和Western blotting测定其效价和特异性,经ELISA检测表明抗血清可与Pep发生特异性免疫反应,经Western blotting试验表明抗血清和多克隆抗体可识别HA特异性条带,其相对分子量为28kD,与预测分子量相符。该抗体的制备为研究HA免疫调控机制提供了有用工具。The immune regulation mechanism of HA gene was studied by using the synthesized HA peptide to prepare the polyclonal antibody. According to primary structure of information about HA in GenBank and the analysis of HA antigen of biological information, three sequence-specific polypeptides were synthesized by fmco solid phase synthesis methods. The purity and molecular weight were determined using HPLC and LC · MS, which showed the purity value reaching at 91.2%, and the molecular weight was 28 kD. The polypeptides were coupled to keyhole limpet hemocyanin （KLH）, and the anti-sera and polyclonal antibody were acquired by immunizing rabbit with them. The titer and specificity of anti-sera and polyclonal antibody were determined by ELISA and Western blotting. The results showed that the an- ti-sera and polyclonal antibody reacted with Pep-KLH and detected a specific band of 28kD, and the size was agreed with the predicted molecular mass. The study provides an important tool to explore the roles of immune regulation mechanism of HA gene.

关 键 词：HA 序列分析 多肽 多克隆抗体 

分 类 号：S811.6[农业科学—畜牧学]