Production of increased titer retrovirus vectors from stable producer cell lines by superinfection and concentration.

Retrovirus vector titers could be increased by more than 50-fold after multiple superinfections of packaging cells with vector virus expressing a complementing host-range envelope glycoprotein. The high level of expression was stable for at least 90 passages in culture. The optimum conditions for density of cell seeding, length of incubation, and temporary storage of virus-containing medium was determined. This vector preparation was further concentrated by 20-fold or more in commercially available ultrafilters, which can be used with standard laboratory centrifuges. Ultrafilters from three different manufactures, with molecular weight cutoffs of 30-1000 kiloDaltons, were tested for concentrating ability and for toxicity of the concentrate on target cells. These methods can be used to produce vector virus containing retrovirus envelope proteins in milliliter volumes at titers in excess of 10(8) c.f.u./ml and in liter volumes at greater than 10(6) c.f.u./ml.