Purification of HPV-HIV VLP Vaccine

A total of 10⁸ transfected 293F cells in a 125 mL Erlenmeyer flask (30 mL culture medium/flask) were collected by centrifugation at 1500 rpm for 5 min and washed twice with PBS. Cell pellets were resuspended in lysis buffer formulated with 1% Triton X-100, protease inhibitor (1:100) (Millipore) and Benzonase (25 U/mL) (Millipore). Cell lysates were clarified with 0.45 µm PVDF syringe filter (Millipore). The HPV:HIV (L1:P18I10 and L1:T20) VLP samples were serially purified using cation exchange (Capto SP ImpRes, GE, Boston, MA, USA), size exclusion (Capto Core 700, GE) and affinity (HiTrap Heparin HP, GE) chromatography. The chromatographic protocols were described in our previous studies [36 (link),37 (link)] and following the manufacturer’s protocol [38 ]. The L1 protein signal in each purification step was characterized by Western blot analysis and probed with anti-HPV16 L1 antibody CAMVIR-1 [39 (link)].