Wang J.,National Vaccine & Serum Institute
Zhonghua shi yan he lin chuang bing du xue za zhi = Zhonghua shiyan he linchuang bingduxue zazhi = Chinese journal of experimental and clinical virology | Year: 2011
To establish the method to detect the cellular immune response of enhanced hepatitis B vaccine and make verification preliminary. Immunized BALB/c mice with enhanced hepatitis B vaccine and detected the IFN-gamma spots forming cells (SFC) of mouse spleen cell by Elispot. Optimized the conditions of the experiment. Cellular immune response between enhanced hepatitis B vaccine and normal hepatitis B vaccine by Elispot were compared. IFN-gamma SFC was higher in 5microg dose than in 2microg dose after immunization with enhanced hepatitis B vaccine and IFN-gamma SFC was declined after immunization 3 weeks ago. IFN-gamma SFC was higher in stimulus by peptide than by protein. Compared to normal hepatitis B vaccine, IFN-gamma SFC was higher in enhanced hepatitis B vaccine. Established the detection method to evaluate the cellular immunity of enhanced hepatitis B vaccine and tested the repeatability.
Liu Y.,National Vaccine & Serum Institute |
Wang X.-X.,National Vaccine & Serum Institute |
Song D.-M.,National Vaccine & Serum Institute |
Wen Z.-H.,National Vaccine & Serum Institute |
And 3 more authors.
Chinese Journal of Biologicals | Year: 2014
Objective To validate the suitability and apply TaqMan MGB probe real-time quantitative PCR for the detection of residual DNA in Vero cells. Methods The TaqMan MGB probe real-time quantitative PCR was validated for specificity, sensitivity, precision and accuracy, and used for detection of residual Vero cell DNA contents in three batches of concentrated, purified and bulk poliovirus, based on which the removal rate of residual Vero cell DNA was calculated. Results The Vero cell genomic DNA long tandem repeats were amplified by the developed TaqMan MGB probe real-time quantitative PCR specifically. Standard curve showed good linearity within a DNA concentration range of 10-1 - 10-6 ng/μl, with a correlation coefficient ( R2 value) of 0. 996 and an amplification efficacy of 93. 54%. The sensitivity of the developed method was 10-6 ng /μl, which was higher than that of dot blot. The coefficients of variation ( CVs ) of determination results of positive control DNA templates at high ( 10-2 ng/μl), moderate ( 10-4 ng /μl ) and low (10-6 ng/ μl) concentrations in intra-assay were 0. 145% ∼ 3. 110%, while those in inter-assay were 0. 624% ∼ 2.359%, respectively. The recovery rates of positive control DNA at various concentrations were 101. 5% ~ 109. 4%, which were within the receptible range. The result of residual Vero cell DNA concentration detected by TaqMan MGB probe real-time quantitative PCR was in line with that by dot blot within the sensitivity range. The total removal rate of residual Vero cell DNA in three batches of inactivated polio vaccine after purification was nearly 100%. Conclusion TaqMan probe realtime quantitative PCR assay showed high specificity, precision, accuracy and sensitivitv, which may be used for in-house quality control of residual Vero cell DNA instead of dot blot hybridization.