Method for large scale production and purification of parvovirus

A method for producing empty inactive or full active parvovirus particles, wherein the parvovirus is H-1PV, said method comprising:

• (a) providing the producer cell line NB-324K;
• (b) growing the cell line under suitable conditions and infecting the cells at a cell density from 2.0 to 5.0×104 cells/cm2 with the parvovirus at a MOI of 0.5 to 2×102 PFU/cells;
• (c) harvesting the cells 2 to 6 days post-infection and obtaining a cell pellet by centrifugation;
• (d) subjecting the resuspended cell pellet to a mechanical, physical or chemical cell lysis method for obtaining a parvovirus containing cell lysate;
• (e) sonicating the cell lysate and subjecting it to DNAse treatment;
• (f) clarifying the DNAse-treated parvovirus harvest by filtration;
• and
• (g1) purifying the parvovirus by two successive density gradient ultracentrifugations, wherein the first gradient is a Iodixanol/PBS step gradient and the second gradient is a Iodixanol/Ringer step gradient or a Iodixanol/Ringer continuous gradient for obtaining full active parvovirus particles in one fraction and empty parvovirus particles in another fraction.