Optimization of poly(ethylene glycol) precipitation of hepatitis A virus used to prepare VAQTA, a highly purified inactivated vaccine.

Abstract

Poly(ethylene glycol) precipitation has been successfully used to concentrate and purify hepatitis A virus from crude lysate preparations for production of VAQTA, a highly purified, formalin-inactivated hepatitis A vaccine. Initial results showed that nucleic acids present in the starting material were problematic for the performance of the poly(ethylene glycol) precipitation step. Extensive experiments were carried out to identify processing conditions suitable for vaccine manufacture which would enhance product yield and improve purity. Results of these studies indicated that the earlier practice of concentrating crude virus-containing lysate using semipermeable membranes led to aggregation of high molecular weight nucleic acids. This aggregated material coprecipitated with the virus during the subsequent poly(ethylene glycol) precipitation step; variable amounts of nucleic acids led to inconsistent virus recovery and product purity. Nuclease treatment of the crude lysate preparations decreased the molecular size of the nucleic acids and significantly reduced their coprecipitation with the virus. Further experiments demonstrated that optimal placement of the nuclease treatment was at the lysate stage followed by a capture step using anion exchange chromatography. These steps combined with optimization of the virus concentration, ionic strength, and pH of the poly(ethylene glycol) precipitation led to effective and selective concentration of the virus which significantly enhanced process reproducibility and control.