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目的了解重组抗原免疫恒河猴的抗体应答情况及其对病毒攻击的保护作用。方法利用大肠杆菌表达的戊型肝炎(HEV)病毒3个不同的片段,分别为Ag1(ORF2C端431aa)、Ag2(ORF2C端128aa片段)、Ag3犤ORF3122aa与ORF26287-6404碱基编码的39aa片段以及C端128aa依次以(Gly)n短臂连接的嵌合抗原犦免疫恒河猴,用ELISA方法检测IgG抗体水平,3周后达到较高水平应答时,用HEV阳性粪便样静脉注射进行攻击,经肝脏病理切片观察、血清IgG、丙氨酸转氨酶(ALT)水平测定及粪便排毒的RT-nPCR检测来评价抗原对病毒攻击的保护作用。结果对照组3只猴中两只发现肝脏组织产生明显的病理性改变,而免疫了抗原的试验组12只猴正常;对照组均在攻毒后3～4周发生ALT异常,为正常值的10～20倍,而试验组有部分猴不同程度轻微的ALT异常,其余正常;试验组免疫抗原后IgG水平迅速上升,滴度可达1∶12800;对照组粪便中病毒RNA的检出在攻毒后7～50d左右,而试验组7～21d内以低水平存在。结论重组的抗原片段诱发机体产生针对HEV的抗体应答,能够抑制HEV感染引起的肝炎症状的产生。
Objective To understand the antibody response of rhesus monkey immunized with recombinant antigen and its protective effect against virus challenge. Methods Three different fragments of HEV virus expressed in Escherichia coli were used to amplify the 39aa fragment encoding Ag1 (ORF2C-terminal 431aa), Ag2 (ORF2C-terminal 128aa), Ag3 犤 ORF3122aa and ORF26287-6404, and C-terminal 128aa was sequentially immunized with chimeric antigen 短 (Gly) n short arm linked to rhesus monkeys. The level of IgG antibody was detected by ELISA. After reaching a higher level of response after 3 weeks, HEV-positive stool- The protective effects of antigens on virus challenge were evaluated by histological observation of liver, determination of serum IgG, alanine aminotransferase (ALT) level and RT-nPCR of excretion detoxification. Results Two of the three monkeys in the control group were found to have obvious pathological changes in the liver tissues. Twelve monkeys in the immunized group were normal. In the control group, abnormal ALT occurred 3 to 4 weeks after challenge 10 ~ 20-fold, while some of the experimental group of mice with varying degrees of mild ALT abnormalities, and the rest of the normal; experimental group immunization antigen IgG levels rose rapidly, the titer up to 1: 12800; control group stool virus RNA detected in the attack After poisoning about 7 ~ 50d, while the experimental group 7 ~ 21d with low levels. Conclusion Recombinant antigens can induce the body to produce antibody response to HEV, and can inhibit the hepatitis symptom caused by HEV infection.