汉坦病毒荧光抗体中和试验方法的建立

Establishment and application of fluorescent antibody virus neutralization test for Hantavirus

目的 荧光抗体病毒中和试验（FAVN）技术应用于汉坦病毒中和抗体的检测的探索与改进。方法采用固定病毒稀释血清的方法，将不同稀释度的血清和固定浓度的病毒经37℃中和1h，然后加入到Vero—E6细胞已长成单层的96孔板上，37℃、5％CO，培养7d，经固定和荧光染色，即可在荧光显微镜下判定结果，根据公式计算血清的中和滴度。结果最合适的细胞浓度为4×10^4～4．5×10^4／孔，病毒血清混合液在37℃中和较好。用此方法检测双价灭活疫苗免疫后30份人血清标本的中和抗体，与空斑减少中和试验结果比较，该方法较敏感、稳定。结论荧光抗体病毒中和试验法具有快速、简便、敏感、稳定的特点。

Objective To establish a fluorescent antibody virus neutralization （FAVN） test for hantavirus （ HV）. Methods With the method of dilution serum and fixed virus, the serum samples were diluted with different concentration, and with virus-liquid were mixed in 96 culture plate hole, 37 ℃for 1 hour. Then the mixture of serum and virus was added to Veto-E6 cells which had grown into a monolayer, and cultivated in the 37℃ , 5% CO2 incubator for 7 days; the 96-well plate was fixed and stained. The results were observed under a fluorescence microscope. The neutralization titer was calculated according to the formula. Results The suitable cell concentrations is 4× 10^4 × 4.5 × 10^4/hole, and the proper temperature for neutralize is 37℃. 30 serum specimens of HFRS were tested by FAVN and PRNT, the FAVN is more sensitive and stable. Conclusion FAVN is easier, faster and more sensitive than the others.