To overcome throughput limitations of existing bionanoparticle purification procedures monoliths are an alternative. The large channels with up to 6 µm in diameter allow the convective transport of virus and virus-like particles (VLPs) in the column. Purification procedure for baculovirus expressed in insect cells and HIV-VLPS expressed in CHO cells have been established. The clarified culture supernatant can be directly loaded on the chromatography column.  With a pre-column functionalised with expoxy groups and a QA-monolith a theoretical yield of infective virus up to 99% could be achieved measured with a plaque forming assay. The virus concentration could be enriched by a factor of 40 making ultrafiltration steps obsolete. The pre-column with epoxy gropus removed phospolipids which lead to fouling to the separation column. We hypothesize that virus adsorbed on phospholipid fouled surfaces fuse to the surface and consequently they cannot be eluted as active viruses. HIC-VLP could be purified also by direct loading on an ion-exchange monolith or a hydrophobic interaction monolith using a hydroxyl functionalized column. In case of HIC the clarified culture supernatant was conditioned with ammonium sulphate. Up to 40% yield has been achieved measured by a laser light particle counting (nanosight) as intact particles. Yield was corroborated by high performance SEC and Westernblots. Depending on the required dose such a purification process would be the standards required for a vaccine. Scale up and scale rules for such a process will be also discussed.<br/>

Purification of viruses and virus-like particles by ion-exchange and hydrophobic interaction monoliths

249th National Meeting (2015)

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