麻腮风水痘联合减毒活疫苗中水痘病毒滴定方法的比较

Comparison of methods for titration of varicella virus in measles,mumps, rubella and varicella combined vaccine, live

目的探讨麻腮风水痘联合减毒活疫苗（measles,mumps,rubella and varicella combined vaccine,live,MMRV）中水痘病毒在人二倍体细胞（MRC-5和2BS）上蚀斑的形成情况,建立不中和法检测MMRV中水痘病毒滴度,并进行验证。方法观察麻疹、腮腺炎和风疹病毒在人二倍体细胞上形成的细胞病变形态,以及风疹病毒对水痘病毒蚀斑形成的影响;通过完全中和、部分中和以及未中和等方法,检测MMRV中水痘病毒的滴度,最终建立不中和法检测水痘病毒滴度,并对方法进行重复性和准确性验证。结果风疹病毒不高于4.0 lg CCID50/孔时,水痘病毒在MRC-5或2BS细胞中产生的蚀斑形态发生微小改变,但蚀斑数量与完全中和风疹病毒后差异无统计学意义（P〉0.5）。采用完全中和、部分中和以及未中和等方法检测MMRV中水痘病毒的滴度差异无统计学意义（P〉0.5）,且不中和法检测水痘疫苗病毒滴度的重复性（RSD为2.3%）和准确性（回收率为97.8%-100%）均较好。结论在人二倍体细胞（2BS或MRC-5）上,不使用抗血清中和直接进行四联疫苗中水痘病毒的滴度检测完全可行,可替代MMRV中经典的使用抗体中和的病毒滴定方法。

Objective To investigate the formation of plaque of varicella virus in measles, mumps, rubella and varicella combined vaccine, live（MMRV） on human diploid cells（MRC-5 and 2BS）, and develop and verify non-neutralizing methods for determination of varicella virus titer in MMRV. Methods The CPEs of measles, mumps and rubella viruses on human diploid cells as well as the effect of rubella virus on plaque formation of varicella virus were observed. The rubella virus titer in MMRV was determined by complete neutralization, partial neutralizaton and non-neutralizing methods,based on which the non-neutralizing method for determination of varicella virus titer was developed, and verified for reproducibility and accuracy. Results When the rubella virus titer was not more than 4. 0 CCID50/ well, the shapes of plaques formed by varicella virus on MRC-5 or 2BS cells showed slight change, while the plaque counts showed no significant difference with those after complete neutralization of rubella virus（P 〉 0. 5）. The varicella virus titers in MMRV determined by complete neutralization, partial neutralizaton and non-neutralizing methods showed no significant difference（P 〉 0. 5）, and the non-neutralizing method showed high reproducibility（RSD was 2. 3%） and accuracy（recovery rate was 97. 8% - 100%）. Conclusion Direct determination of varicella virus in MMRV on MRC-5 and 2BS cells without neutralizing with antisera was feasible, which might be used instead of the classical titration methods using neutralizing antibody.