title: Asymmetric flow field flow fractionation methods for virus purification.
abstract: Detailed biochemical and biophysical characterization of viruses requires viral preparations of high quantity and purity. The optimization of virus production and purification is an essential, but laborious and time-consuming process. Asymmetric flow field flow fractionation (AF4) is an attractive alternative method for virus purification because it is a rapid and gentle separation method that should preserve viral infectivity. Here we optimized the AF4 conditions to be used for purification of a model virus, bacteriophage PRD1, from various types of starting materials. Our results show that AF4 is well suited for PRD1 purification as monitored by virus recovery and specific infectivity. Short analysis time and high sample loads enabled us to use AF4 for preparative scale purification of PRD1. Furthermore, we show that AF4 enables the rapid real-time analysis of progeny virus production in infected cells.
authors.affiliation: Department of Biosciences and Institute of Biotechnology, Viikinkaari 9, University of Helsinki, FIN-00014 Helsinki, Finland.
authors.forename: Katri
authors.initials: K
authors.lastname: Eskelin
authors.affiliation: Department of Biosciences and Institute of Biotechnology, Viikinkaari 9, University of Helsinki, FIN-00014 Helsinki, Finland.
authors.forename: Mirka
authors.initials: M
authors.lastname: Lampi
authors.affiliation: Postnova Analytics, Max-Planck-Str. 14, 86899 Landsberg, Germany.
authors.forename: Florian
authors.initials: F
authors.lastname: Meier
authors.affiliation: Postnova Analytics, Max-Planck-Str. 14, 86899 Landsberg, Germany.
authors.forename: Evelin
authors.initials: E
authors.lastname: Moldenhauer
authors.affiliation: Department of Biosciences and Institute of Biotechnology, Viikinkaari 9, University of Helsinki, FIN-00014 Helsinki, Finland.
authors.forename: Dennis H
authors.initials: DH
authors.lastname: Bamford
authors.affiliation: Department of Biosciences and Institute of Biotechnology, Viikinkaari 9, University of Helsinki, FIN-00014 Helsinki, Finland. Electronic address: hanna.oksanen@helsinki.fi.
authors.forename: Hanna M
authors.initials: HM
authors.lastname: Oksanen
mesh_terms: D025622:Bacteriophage PRD1 / Q000302:isolation & purification; D031521:Fractionation, Field Flow / Q000379:methods; D012486:Salmonella typhimurium / Q000821:virology; D014764:Viral Proteins / Q000032:analysis; D014771:Virion / Q000302:isolation & purification; D014780:Viruses / Q000302:isolation & purification*
chemical_list: D014764:Viral Proteins
keywords: Bacteriophage PRD1; Icosahedral virus; Membrane virus; Monolithic chromatography; Real-time analysis of progeny virus production; Ultracentrifugation