重组狂犬病病毒抗体中和试验方法的建立

Establishment of a recombinant rabies virus antibody neutralization test method

为了构建一种使用重组狂犬病病毒的抗体中和试验方法(FAVN-GFP),将8种兽用狂犬病灭活疫苗(含标准参考疫苗)分别5倍梯度稀释4个稀释度后免疫小鼠,分别在首次免疫后的第14、28天采集小鼠血清,以表达绿色荧光蛋白的重组狂犬病病毒作为抗原,使用FAVN-GFP方法测定中和抗体效价,并与中国兽药典中的经典NIH方法进行对比分析。结果显示,首次免疫后第28天1∶25稀释组对8种疫苗的检测结果与NIH法的结果在斜率/差分上显著相关,证明在该条件下两种方法具有相同的变化趋势。结果表明,本实验所建立的FAVN-GFP法可以在兽用狂犬病灭活疫苗的效力评价中替代现有的NIH方法,能够大量减少实验动物的使用,更加安全、准确和经济。

In order to construct a recombinant rabies virus antibody neutralization test method(FAVN-GFP),eight veterinary inactivated rabies vaccines(including standard reference vaccines)were diluted to four dilutions by five-fold serial dilution.Then mice were immunized with these diluted vaccines.The mouse serum was collected on the 14th and 28th days after the first immunization.Using the recombinant rabies virus expressing green fluorescent protein as antigen,the neutralizing antibody titer was determined by FAVN-GFP method.This serological test were compared with the National Institutes of Health(NIH)method in the Chinese Veterinary Pharmacopoeia.The results showed that the two methods were significantly correlated in slope/differential for eight vaccines at 1:25 dilution on 28th day after the first immunization,which proved that the two methods had the same change trend under this condition.In conclusion,the FAVN-GFP method established in this laboratory can be applied in the potency evaluation of veterinary inactivated rabies vaccine,and can replaced the existing NIH method,which will greatly reduce the use of laboratory animals while being safer,more accurate and economical.