Cellular Impedance-Based Survival Kinetics Assay to Assess Virus Survival
Transcrição
To assess the influenza A virus survival kinetics, add sodium chloride to a final concentration of 35 grams per liter in distilled water, and add 900 microliters of the resulting saline solution to 2-milliliter cryotubes. Add 100 microliters viral stock to each cryotube, and place the tubes in the 35-degree Celsius incubator, for the appropriate experimental time period.
The day before the end of the incubation, seed 3 x 104 freshly-split MDCK cells in 100 microliters of virus propagation medium per well, in a 16-well microtiter plate, and repeat steps allowing measurement of the background impedance and uniform distribution of the cells onto the bottom of the wells. Then, grow the cells for 24 hours at 37 degrees Celsius and 5% carbon dioxide.
The next day, wash the cells twice, then, infect the cells with 100 microliters of the saline-distilled virus, diluted 10 times in fresh culture medium, as demonstrated, and monitor the cell impedance every 15 minutes for at least 100 hours.
