Purification of foamy viral particles

Abstract

Foamy viruses are non-pathogenic retroviruses and represent a tool for vector development. For gene therapy applications and for analyses of viral protein composition infectious particles need to be purified, which has been difficult for foamy viruses in the past. Here, we describe a novel, simple, and fast purification method for prototype foamy viruses with high purity using size exclusion and affinity chromatography. More than 99,9% of the contaminating proteins were removed. The purified viruses were used to determine the amount of the incorporated Pol protein relative to Gag. The determined Gag to Pol PR-RT ratio of 30:1 confirmed previous studies suggesting FV virions encapsidate fewer number of Pol molecules than orthoretroviruses.

FV Vectors as Alternative Gene Vehicles for Gene Transfer in HSCs

“…Foamy viruses have the largest genome among retroviruses and as a result, FV vectors have enough space to accommodate a little over 9 kb of exogenous DNA [44]. and ultracentrifugation [51]. Both purification approaches are performed in closed systems compliant with cGMP and yield significantly pure preparations.…”

“…These modifications rendered FV vectors as efficient as lentiviral vectors producing 10 × 10 7 TU/mL of crude supernatant [44]. Finally, FV supernatants can be concentrated and purified in closed systems using affinity chromatography with POROS-Heparin columns, followed by Tangential Flow Filtration and ultracentrifugation [50] or size exclusion chromatography using CaptoCore columns followed by sepharose-heparin affinity chromatography and ultracentrifugation [51]. Both purification approaches are performed in closed systems compliant with cGMP and yield significantly pure preparations.…”