禽源糖原合成激酶蛋白单克隆抗体的制备及应用  

Preparation and Application of Monoclonal Antibody Against Avian Glycogen Synthase Kinase Protein

作　　者：翁贝 崔洲昊 邵红霞 秦爱建[1,2,3] 钱琨 WENG Bei;CUI Zhouhao;SHAO Hongxia;QIN Aijian;QIAN Kun(Key Laboratory for Avian Preventive Medicine,Ministry of Education,Yangzhou University,Yangzhou,Jiangsu 225009;Key Laboratory of Jiangsu Preventive Veterinary Medicine,Yangzhou University,Yangzhou,Jiangsu 225009;Jiangsu Co-innovation Center for the Prevention and Control of Important Animal Infectious Diseases and Zoonoses,Yangzhou,Jiangsu 225009)

机构地区：[1]扬州大学教育部禽类预防医学重点实验室,江苏扬州225009 [2]扬州大学江苏省动物预防医学重点实验室,江苏扬州225009 [3]江苏高校动物重要疫病与人兽共患病防控协同创新中心,江苏扬州225009

出　　处：《中国家禽》2024年第10期81-88,共8页China Poultry

基　　金：国家自然科学基金项目(32072839、32272970);江苏高校“青蓝工程”资助项目(苏教师函[2021]11号)。

摘　　要：为制备针对家禽糖原合成酶激酶3β(GSK-3β)的单克隆抗体以及了解不同细胞中GSK-3β蛋白的基础表达量,试验利用原核表达纯化的GSK-3β蛋白免疫BALB/c小鼠,真核表达GSK-3β蛋白间接免疫荧光筛选杂交瘤阳性细胞,并利用单克隆抗体检测不同禽源细胞(LMH细胞、DF-1细胞、CEF细胞)中GSK-3β蛋白表达情况。结果显示:共获得3株单克隆抗体细胞株,分别命名为4C1、5B11和8G9,其中4C1和5B11仅能结合禽源细胞中的GSK-3β,而8G9和禽源以及人源的GSK-3β都能反应;单克隆抗体4C1介导的Western bolt显示在LMH细胞中GSK-3β蛋白丰度相对最高,而RT-qPCR显示其中LMH细胞中GSK-3β基因转录水平较低。研究成功制备了禽源GSK-3β的单克隆抗体,为深入研究家禽GSK-3β的生物学功能提供良好的生物材料。In order to prepare monoclonal antibody against poultry glycogen synthetase kinase 3β(GSK-3β)and understand the basal expression of GSK-3βprotein in different cells,BALB/c mice were immunologized with prokaryotic expression purified GSK-3βprotein,and hybridoma-positive cells were screened by indirect immunofluorescence of eukaryotic expression GSK-3βprotein,and the expression of GSK-3βprotein in different avian derived cells(LMH cell,DF-1 cell,CEF cell)was detected by monoclonal antibodies.Three monoclonal antibody cell lines were screened,named 4C1,5B11 and 8G9,among which 4C1 and 5B11 could only bind GSK-3βin avian cells,while 8G9 could react with GSK-3βfrom avian and human sources.The monoclonal antibody 4C1-mediated Western bolt showed that the abundance of GSK-3βprotein was the highest in LMH cell,while RT-qPCR showed that the transcription level of GSK-3βgene was lower in LMH cell.The monoclonal antibodiesy against poultry GSK-3βwas successfully prepared,which provided good biological materials for in-depth study of biological function of GSK-3βin poultry.

关 键 词：禽 GSK-3Β 原核表达 单克隆抗体 应用 

分 类 号：S855.3[农业科学—临床兽医学]