不同ELISA检测方法在检测出壳雏鸡禽白血病病毒感染中的应用  

Application of Different ELISA Detection Methods in Detecting Avian Leukosis Virus Infection in Chickens

作　　者：崔慧珍 张亚文 王一新[1] 常爽[1] 赵鹏[1] CUI Huizhen;ZHANG Yawen;WANG Yixin;CHANG Shuang;ZHAO Peng(College of Animal Science and Veterinary Medicine,Shandong Agricultural University,Tai'an 271018,China)

机构地区：[1]山东农业大学动物科技学院,山东泰安271018

出　　处：《中国兽医杂志》2024年第4期27-34,共8页Chinese Journal of Veterinary Medicine

基　　金：山东省重点研发计划(乡村振兴科技创新提振行动计划)(2022TZXD0019)。

摘　　要：通过雏鸡胎粪中群特异性衣壳蛋白27(p27)抗原检测以判定禽白血病病毒(ALV)垂直传播是实施禽白血病净化的关键步骤,为对比不同ALV-p27抗原ELISA试剂盒对我国四类广泛流行的亚群ALV垂直传播的检测效果,本试验设置ALV-A、ALV-B、ALV-J和ALV-K感染组,以无菌生理盐水作为对照组,在8胚龄时以卵黄囊接种的方式感染SPF鸡胚,建立鸡胚携带ALV-A、ALV-B、ALV-J和ALV-K的垂直感染模型。采集各组雏鸡胎粪样品和血浆样品,死亡鸡胚肝脏样品,死亡雏鸡胎粪样品、血浆样品和肝脏样品,其中血浆和肝脏样品需接种鸡胚成纤维细胞系(DF-1细胞)进行病毒分离,各样品以4家公司提供的ALV-p27抗原ELISA试剂盒进行检测,并以实时荧光定量PCR(RT-qPCR)进行平行检测,比较阳性检出率和灵敏度。结果显示,4家公司的ALV-p27抗原ELISA试剂盒对于同一样品的阴阳性判定结果基本一致,但灵敏度存在差异;胎粪样品直接ELISA检测不能将ALV阳性鸡只全部检出,相对于胎粪样品,血浆和肝脏病毒分离样品ELISA检测更具有灵敏度优势。本试验所建立的对比方法不仅有助于评估4家公司ELISA试剂盒的灵敏度,还进一步评估了胎粪样品直接ELISA检测与血浆和肝脏病毒分离样品ELISA检测2种评估方法的灵敏度,为科学选择灵敏特异的ALV检测方法提供了良好评价模型。Determining the vertical transmission of avian leukosis virus(ALV)through the detection of group-specific protein 27(p27)in chick meconium is a key step in implementing ALV eradication.To compare the detection effects of different ALV-p27 antigen ELISA kits on four widely prevalent subgroups of ALV in China vertical transmission,this study established ALV-A,ALV-B,ALV-J,and ALV-K infection groups,with sterile physiological saline as the control group.SPF chicken embryos were infected by inoculating into the yolk sac at embryonic day 8 to establish vertical infection models carrying ALV-A,ALV-B,ALV-J,and ALV-K.Meconium and plasma samples of each group,liver samples of dead chicken embryos,and meconium,plasma,and liver samples of dead chicks were collected.Plasma and liver samples were inoculated onto Douglas foster-1(DF-1)cells for virus isolation.Each sample was tested using ALV-p27 antigen ELISA kits provided by four companies,and parallel testing was conducted using real-time quantitative PCR(RT-qPCR)to compare the positive detection rates and sensitivity.The results showed that the ALV-p27 antigen ELISA kits from the four companies had generally consistent positive and negative results for the same samples,but there were differences in sensitivity.Direct ELISA detection of meconium samples could not detect all ALV-positive chickens.Compared with meconium samples,ELISA detection of plasma and liver virus isolation samples had greater sensitivity.The comparative method established in this study not only helps evaluate the sensitivity of ELISA kits from four companies but also further evaluates the sensitivity of two evaluation methods:direct ELISA detection of meconium samples and ELISA detection of plasma and liver virus isolation samples,providing a good evaluation model for scientifically selecting sensitive and specific ALV detection methods.

关 键 词：禽白血病病毒 胎粪 P27抗原 ELISA 病毒分离 

分 类 号：S855.3[农业科学—临床兽医学]