Polybrene Infection / Transfection Reagent Protocol

Retroviral Infection

1. Recombinant retroviral stocks are prepared by adding 5 mL of growth medium with 5% serum to a near confluent monolayer of transfected retroviral packaging cells in a 100 mm plate. After 24 hours the medium is removed and filtered through a 0.45 µm filter.
2. Cells to be infected with this recombinant retroviral stock are plated at 500,000 cells per 100 mm plate in 10 mL of complete medium.
3. Remove the growth medium from the cells 24 hours later. Infect cells with 2 mL of the viral supernatant (or a dilution of the virus stock into 2 mL) in the presence of 5 µg to 10 µg of polybrene per mL (final concentration). Incubate cells for 3 to 6 hours at 37°C.
4. Add 8 mL of complete medium. Three days after infection, split the cells 1:5 into selection medium.

References

1. Toyoshima, K. and Vogt, P.K., Virology 1969; 38:414–26
2. Coelen, J.R., Jose, D.G. and May, J.T. Arch. Virol. 1983; 75:307–11