别藻蓝蛋白标记抗鸡IgG荧光抗抗体的高效制备与鉴定  

Preparation and identification of allophycocyanin-labeled anti-chicken IgG fluorescence antibody

作　　者：朱丽萍[1,2,3] 颜世敢[2,3,4] 李雁冰[2] 张玉忠[3] 

机构地区：[1]山东轻工业学院食品与生物工程学院,山东济南250353 [2]中国农业科学院哈尔滨兽医研究所兽医生物技术国家重点实验室,黑龙江哈尔滨150001 [3]山东大学微生物技术国家重点实验室,山东济南250100 [4]山东省农业科学院畜牧兽医研究所,山东省动物疫病防控与繁育重点实验室,山东济南250100

出　　处：《江苏农业学报》2011年第1期110-115,共6页Jiangsu Journal of Agricultural Sciences

基　　金：兽医生物技术国家重点实验室开放基金项目(SKLVBF200909);微生物技术国家重点实验室开放基金项目(SKLMTF200818)

摘　　要：将别藻蓝蛋白(Allophycocyanin,APC)与抗鸡IgG抗体交联制备荧光抗抗体,用于鸡疫病的间接荧光检测。首先采用羟基磷灰石层析和阴离子交换层析法高效制备APC,然后利用蛋白质交联技术高效制备荧光抗抗体。分别用摩尔比50∶1、100∶1的交联剂3-(2-吡啶二巯基)丙酸N-羟基琥珀酰亚胺酯(SPDP)将APC、抗鸡IgG抗体衍生,衍生物以摩尔比1∶1液相交联,经高压液相色谱(HPLC)纯化制备出APC标记的抗鸡IgG荧光抗抗体,最后对制备的荧光抗抗体从荧光特性、吸收光谱特性、抗体活性、稳定性及纯度等方面进行鉴定。结果表明,APC标记的抗鸡IgG荧光抗抗体交联效率达95%以上,纯度达电泳纯,红色荧光明亮,抗体活性高,4℃保存90 d荧光亮度和抗体活性稳定。研制的APC标记的抗鸡IgG荧光抗抗体可作为通用荧光抗抗体用于鸡传染性疾病的间接免疫荧光检测。Allophycocyanin（APC） was firsly purified by hydroxylapatite and anion exchange chromatography,then APC and anti-chicken IgG antibody were chemically modified by crosslinker SPDP at a ratio of 50 and 100 to 1.APC ramification was conjugated with anti-chicken IgG antibody at a ratio of 1 to 1.Finally APC-labeled anti-chicken IgG fluorescence antibody was prepared after purification by HPLC.Its absorption and fluorescence spectra,antibody bioreactivity,electrophoresis and stability were determined.Results revealed the conjugation efficiency ratio reached as high as 95% and the APC-labeled anti-chicken IgG fluorescence antibody was pure,highly fluorescent,bioreactive and stable.The prepared APC-labeled anti-chicken IgG fluorescence antibody was suitable for indirect immunofluorescence detection for chicken infectious diseases.

关 键 词：别藻蓝蛋白 蛋白质标记 荧光抗抗体 

分 类 号：S858.31[农业科学—临床兽医学]