title: Purification of infective baculoviruses by monoliths.
abstract: A chromatographic process based on monoliths for purification of infective baculovirus without prior concentration step has been established. Baculovirus produced in Spodoptera frugiperda cells (Sf-9) were harvested by centrifugation, filtered through 0.8 μm filters and directly loaded onto radial 1 mL anion exchange monoliths with a channel size of 1.5-2.0 μm operated at a volumetric flow rate of one bed volume per minute. Optional an epoxy monolith was used as pre-column to reduce interfering compounds and substances influencing the capacity of anion exchange monoliths for baculovirus infectious virus could be eluted with a step gradient at salt concentrations of 440 mM NaCl. Recovery of infectious virus was highly influenced by composition and age of supernatant and ranged from 20 to >99% active baculovirus. Total protein content could be reduced to 1-8% and DNA content to 38-48% in main virus fraction. Infective virus could be 52-fold concentrated within 20.5h and simultaneously an 82-fold volume reduction was possible when loading 1150 mL (2.1×10(8) pfu/mL) onto 1 mL scale support.
authors.affiliation: Department of Biotechnology, University of Natural Resources and Life Sciences Vienna, Vienna, Austria.
authors.forename: Petra
authors.initials: P
authors.lastname: Gerster
authors.forename: Eva-Maria
authors.initials: EM
authors.lastname: Kopecky
authors.forename: Nikolaus
authors.initials: N
authors.lastname: Hammerschmidt
authors.forename: Miriam
authors.initials: M
authors.lastname: Klausberger
authors.forename: Florian
authors.initials: F
authors.lastname: Krammer
authors.forename: Reingard
authors.initials: R
authors.lastname: Grabherr
authors.forename: Christa
authors.initials: C
authors.lastname: Mersich
authors.forename: Lidija
authors.initials: L
authors.lastname: Urbas
authors.forename: Petra
authors.initials: P
authors.lastname: Kramberger
authors.forename: Tina
authors.initials: T
authors.lastname: Paril
authors.forename: Matthias
authors.initials: M
authors.lastname: Schreiner
authors.forename: Katharina
authors.initials: K
authors.lastname: Nöbauer
authors.forename: Ebrahim
authors.initials: E
authors.lastname: Razzazi-Fazeli
authors.forename: Alois
authors.initials: A
authors.lastname: Jungbauer
mesh_terms: D000818:Animals; D000838:Anions / Q000737:chemistry; D016367:Baculoviridae / Q000302:isolation & purification*; D015153:Blotting, Western; D018929:Cell Culture Techniques; D002852:Chromatography, Ion Exchange / Q000295:instrumentation / Q000379:methods*; D004591:Electrophoresis, Polyacrylamide Gel; D008055:Lipids; D010316:Particle Size; D015203:Reproducibility of Results; D061987:Sf9 Cells / Q000821:virology*
chemical_list: D000838:Anions; D008055:Lipids