Propagation and Harvesting of Oncolytic Herpes Simplex Virus in Mammalian Cells

Take a culture of mammalian cells in a growth medium. Remove the medium and wash with a buffer.
Add the oncolytic herpes simplex virus suspension and agitate for even distribution.
During incubation, the viral glycoproteins bind to the host receptors, enabling adsorption.
Remove the unadsorbed viruses and add the medium.
Upon incubation, the viral envelope fuses with the host membrane, releasing the nucleocapsid into the cytoplasm.
The nucleocapsid delivers the viral genome into the nucleus, where the host machinery is hijacked to produce progeny viruses.
The rising viral load causes structural damage to the cells.
Collect the conditioned medium and scrape to detach the cells. Add the conditioned medium, transfer them to a tube, and spin down the cells.
Discard the supernatant. Resuspend in a buffer and the conditioned medium to maintain virus stability.
Snap-freeze the suspension and store at ultra-low temperatures to preserve the virus structure.

Begin by washing the T-150 square centimeter flasks with two times high glucose DPBS supplemented with 1% IFCS. Aspirate the DPBS and add seven milliliters of virus inoculum per flask.

Gently rock the flasks for five minutes and incubate at 37 degr