Heparin-affinity chromatography is a generic purification platform for chimeric gag VLPs displaying different viral surface antigens

Increased efficacy of influenza virus vaccine candidate through display of recombinant neuraminidase on virus like particles

“…Gag-based VLPs displaying the N2 were expressed using the BEVS and the T.ni insect cell line, Tnms42 ( 35 ). NA-VLPs were then purified via a platform downstream procedure based on a Capto-Core 700™ flow-through chromatography followed by a Capto-Heparin™ affinity chromatography ( 26 ). The flow-through material from the Capto-Core 700 ™ was loaded onto the heparin column to separate the gag-VLPs from baculovirus particles and other extracellular vesicles.…”

“…This is consistent with reports showing that NAs express at lower levels than HAs ( 30 ). Moreover, NA-VLPs underwent a two-step chromatography purification process, which has previously been successful in producing high-purity VLPs ( 26 ). Semi-purified VLP material has been used up to this point, which may have caused an overestimation of the antigen’s quantification because extracellular vesicles (EVs) and BVs share the same cell membrane and exhibit proteins on the surfaces that are similar to those of VLPs, such as NA or HA.…”

“…The protein was concentrated using an Amicon Ultracel-15 centrifugal filter with a cut-off of 30 kDa (Millipore) and the buffer was exchanged to phosphate buffered saline (PBS) pH 7.4. For the VLPs, a platform purification method ( 26 ) was combined with ultracentrifugation to concentrate the material. The clarified supernatant was filtered with a 0.8 µm sterile polyvinylidene difluoride (PVDF) syringe filter (Thermo Fisher Scientific).…”

“…N2 was chosen because of the high diversity of H3N2 viruses in the human population and the resulting typically low vaccine effectiveness ( 25 ). These VLPs were expressed and purified using a platform downstream process based on Capto-Core 700™ flow-through chromatography followed by Capto-Heparin™ affinity chromatography ( 26 ) combined with ultracentrifugation and were then tested in the mouse model. VLPs displaying NA as described here could, in the future, be added to seasonal influenza vaccines to increase immune responses to NA.…”