Amplification of P3 stock
1. Prepare cells:
a. It is also recommended to follow the growth of the culture. The cell concentration should be monitored (and maintained <5x10^6/ml).
b. The culture should be monitored daily and harvested 3 days post-growth cessation or when culture viability has reached approx. 50%.
2. Cultures are harvested at 3 to 4 days post-infection.
a. Important: at times of infection add heat-inactivated fetal bovine serum (FBS) at 5% (v/v) to the culture for protection of the virus.
b. Heat inactivation of FBS
3. Virus preps are centrifuged at 2000 x g for 30 min to clarify them.
4. Store at 4°C in the dark (put the bottle in light-proof bags).
5. The final titer must be greater than 5 x 10^7 pfu/ml.
6. Containers for the storage of virus stocks.