PRODUCTION OF INCREASED TITER RETROVIRUS VECTORS FROM STABLE PRODUCERCELL-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 express ing a complementing host-range envelope glycoprotein. The high level o f expression was stable for at least 90 passages in culture. The optim um conditions for density of cell seeding, length of incubation, and t emporary storage of virus-containing medium was determined. This vecto r preparation was further concentrated by 20-fold or more in commercia lly available ultrafilters, which can be used with standard laboratory centrifuges. Ultrafilters from three different manufactures, with mol ecular weight cutoffs of 30-1000 kiloDaltons, were tested for concentr ating ability and for toxicity of the concentrate on target cells, The se methods can be used to produce vector virus containing retrovirus e nvelope 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.