Objective: To develop and verify a fluorescent quant. reverse transcription-polymerase chain reaction (RT-PCR) assay for infectious titer of live attenuated mumps vaccine.Methods: Primers and Taq Man fluorescent probe were designed specific to the conserved region of hemagglutinin (H) gene of live attenuated mumps vaccine virus strain S79.The final product prepared with strain S79, with a lot Number of 05008, was served as a referenceVero cells were inoculated with the reference or test sample, then broken by hypoosmotic method combined with freeze-thawing method, of which the supernatant was collected and determined by fluorescent quant. RT-PCR.The time for virus infection was optimized, and the developed method was verified for specificity, precision and accuracy.Results: The optimal time for virus infection was 18 h.The developed fluorescent quant. RT-PCR assay was specific to live attenuated mumps vaccine, while no amplification curves were observed in the vaccine after inactivation, varicella virus, rabies virus, measles virus, rebulla virus or Vero cells.The relative standard deviations (RSDs) of six groups of data on samples at four concentrations (1, 1:5, 1:52 and 1:53) were less than 5%.The R2 values of regression equation of standard curves of samples at four concentrations (1, 1:5, 1:52 and 1:53) determined by various personnel on various dates were more than 0.97, with RSDs of less than 5%.The difference between determination results of 11 batches of live attenuated mumps vaccine by the developed method and by CPE method was not more than 0.2 Lg CCID50/mL, which showed no significant difference (P>0.05).Conclusion: The developed fluorescent quant. RT-PCR assay for infectious titer of live attenuated mumps vaccine showed high specificity, precision and accuracy, which was rapid, simple, and suitable for the inhouse quality control during production