Production and concentration of pseudotyped HIV-1-based gene transfer vectors.
Strategies to generate highly concentrated HIV-1 vector pseudotypes involving different envelope (Env) proteins including the vesicular stomatitis virus (VSV) G glycoprotein, the Moloney murine leukemia virus (MLV) 4070A amphotropic Env and the rabies G glycoprotein were established. Virus stocks were prepared by transient transfection using standard cell culture media or serum-free media. Such stocks were concentrated 50- to 300-fold by ultracentrifugation or by ultrafiltration using Centricon Plus-80 units yielding titers of up to 109transducing units per milliliter. There was no loss in titer with any of the pseudotypes tested. Thus, like lentiviral vectors pseudotyped with VSV-G, HIV-1-based vectors pseudotyped with the MLV 4070A amphotropic Env and the rabies G glycoprotein resist inactivation during concentration. This opens up the possibility to generate highly concentrated HIV-1 vector stocks carrying alternative Env proteins on a large scale.
Reiser
J
J
Developmental and Metabolic Neurology Branch, National Institute of Neurological Disorders and Stroke, National Institutes of Health, Bethesda, MD, USA.
D002460:Cell Line; D018014:Gene Transfer Techniques; D015316:Genetic Therapy / Q000379:methods*; D005822:Genetic Vectors / Q000235:genetics / Q000302:isolation & purification*; D015497:HIV-1 / Q000235:genetics*; D006801:Humans; D014759:Viral Envelope Proteins / Q000235:genetics*; D014760:Viral Fusion Proteins / Q000235:genetics
D014759:Viral Envelope Proteins; D014760:Viral Fusion Proteins