title: Purification of Vero cell derived live replication deficient influenza A and B virus by ion exchange monolith chromatography.
abstract: We explored the possibilities for purification of various ΔNS1 live, replication deficient influenza viruses on ion exchange methacrylate monoliths. Influenza A ΔNS1-H1N1, ΔNS1-H3N2, ΔNS1-H5N1 and ΔNS1-influenza B viruses were propagated in Vero cells and concentrated by tangential flow filtration. All four virus strains adsorbed well to CIM QA and CIM DEAE anion exchangers, with CIM QA producing higher recoveries than CIM DEAE. ΔNS1-influenza A viruses adsorbed well also to CIM SO3 cation exchanger at the same pH, while ΔNS1-influenza B virus adsorption to CIM SO3 was not complete. Dynamic binding capacity (DBC) for CIM QA, DEAE and SO3 methacrylate monoliths for influenza A ΔNS1-H1N1 virus were 1.9E+10 TCID50/ml, 1.0E+10 TCID50/ml and 8.9E+08 TCID50/ml, respectively. Purification of ΔNS1 viruses on CIM QA was scaled up and reproducibility was confirmed. Yields of infectious virus on CIM QA were between 70.8±32.3% and 87±30.8%. Total protein removal varied from 93.3±0.4% to 98.6±0.2% and host cell DNA removal efficiency was ranging from 76.4% to 99.9% and strongly depended on pretreatment with deoxyribonuclease.
authors.affiliation: BIA Separations, Mirce 21, SI-5270 Ajdovščina, Slovenia.
authors.forename: Marko
authors.initials: M
authors.lastname: Banjac
authors.affiliation: AVIR Green Hills Biotechnology AG, Forsthausgasse 11, A-1200 Vienna, Austria.
authors.forename: Elisabeth
authors.initials: E
authors.lastname: Roethl
authors.affiliation: AVIR Green Hills Biotechnology AG, Forsthausgasse 11, A-1200 Vienna, Austria.
authors.forename: Franz
authors.initials: F
authors.lastname: Gelhart
authors.affiliation: BIA Separations, Mirce 21, SI-5270 Ajdovščina, Slovenia.
authors.forename: Petra
authors.initials: P
authors.lastname: Kramberger
authors.affiliation: BIA Separations, Mirce 21, SI-5270 Ajdovščina, Slovenia.
authors.forename: Barbara Lah
authors.initials: BL
authors.lastname: Jarc
authors.affiliation: BIA Separations, Mirce 21, SI-5270 Ajdovščina, Slovenia.
authors.forename: Marko
authors.initials: M
authors.lastname: Jarc
authors.affiliation: BIA Separations, Mirce 21, SI-5270 Ajdovščina, Slovenia.
authors.forename: Aleš
authors.initials: A
authors.lastname: Strancar
authors.affiliation: AVIR Green Hills Biotechnology AG, Forsthausgasse 11, A-1200 Vienna, Austria.
authors.forename: Thomas
authors.initials: T
authors.lastname: Muster
authors.affiliation: BIA Separations, Mirce 21, SI-5270 Ajdovščina, Slovenia. Electronic address: matjaz.peterka@monoliths.com.
authors.forename: Matjaž
authors.initials: M
authors.lastname: Peterka
mesh_terms: D000327:Adsorption; D000818:Animals; D002522:Chlorocebus aethiops; D002852:Chromatography, Ion Exchange / Q000379:methods*; D005374:Filtration; D053118:Influenza A Virus, H1N1 Subtype / Q000302:isolation & purification* / Q000502:physiology; D053122:Influenza A Virus, H3N2 Subtype / Q000302:isolation & purification* / Q000502:physiology; D053124:Influenza A Virus, H5N1 Subtype / Q000302:isolation & purification* / Q000502:physiology; D009981:Influenza B virus / Q000302:isolation & purification* / Q000502:physiology; D014709:Vero Cells; D014776:Virus Cultivation; D014779:Virus Replication
keywords: Chromatography; Influenza viruses; Monoliths; Purification; Vaccines