A simplified ATP method for the rapid control of cell viability in a freeze-dried BCG vaccine

作者：Ugarova, Natalia N.; Lomakina, Galina Yu.; Modestova, Yulia; Chernikov, Sergey V.; Vinokurova, Natalia V.; Otrashevskaya, Elena V.; Gorbachev, Vyacheslav Y.

来源：Journal of Microbiological Methods, 2016, 130: 48-53.

DOI：10.1016/j.mimet.2016.08.027

• ATP assay
• BCG viability
• Colony forming units
• Freeze-dried BCG vaccine
• Tuberculosis

We propose a simple and cost-effective ATP method for controlling the specific activity of a freeze-dried BCG vaccine. A freeze-dried BCG vaccine is reconstituted with 1 ml saline and incubated for 15 min at room temperature and then for 1 hat 37 degrees C. The vaccine is then treated with apyrase to remove extracellular ATP. After that, the cells are lysed with DMSO and the ATP content in the lysate is measured by the bioluminescence method. To implement the method, we developed a kit that requires no time-consuming preparation before the analysis. We demonstrated the linear relationship between the experimental values of the specific activity (10(6) CFU/mg) and intracellular ATP content (ATP, pmol/mg) for different batches of the studied BCG vaccines; the proportionality coefficient was K = 0.36 +/- 0.02. We proposed a formula for calculating the specific activity from the measured content of intracellular ATP (ATP, pmol/mg). The comparison of the measured and calculated values of the specific activity (10(6) CFU/mg) shows that these values are similar; their differences fall within the allowable range of deviations for the specific activity values of the BCG vaccine.

• 出版日期2016-11