HIV-1NL4-3 stocks were made by transfection of HEK293T cells with a proviral DNA plasmid (39 (link), 40 (link)). The medium was replaced at 24 h posttransfection, and HIV-1 was harvested at 72 h posttransfection. All virus stocks were quantified by a CAp24 enzyme-linked immunosorbent assay (ELISA) and stored in small aliquots at −80°C as previously described (18 (link)).
VLPs that convey NB-ZSG or ZSG were produced as previously described (21 (link)). Briefly, 10 μg of the pSRS11-SF-γC vector (25 (link)) expressing NB-ZSG or ZSG and 2 μg of a plasmid expressing Gag-Pol were transfected into Phoenix-Ampho cells (41 (link)) with X-tremeGENE HP DNA transfection reagent (Roche Diagnostics GmbH, Mannheim, Germany) in accordance with the manufacturer’s protocol. The cell medium was replaced at 24 h posttransfection. VLPs were harvested by filtration with a sterile 0.22-μm syringe filter (Sartorius Stedim Biotech GmbH, Göttingen, Germany) at 48 h and 72 h posttransfection.
VLPs that convey NB-ZSG or ZSG were produced as previously described (21 (link)). Briefly, 10 μg of the pSRS11-SF-γC vector (25 (link)) expressing NB-ZSG or ZSG and 2 μg of a plasmid expressing Gag-Pol were transfected into Phoenix-Ampho cells (41 (link)) with X-tremeGENE HP DNA transfection reagent (Roche Diagnostics GmbH, Mannheim, Germany) in accordance with the manufacturer’s protocol. The cell medium was replaced at 24 h posttransfection. VLPs were harvested by filtration with a sterile 0.22-μm syringe filter (Sartorius Stedim Biotech GmbH, Göttingen, Germany) at 48 h and 72 h posttransfection.
Free full text: Click here
