title: Purification of a recombinant baculovirus of Autographa californica M nucleopolyhedrovirus by ion exchange membrane chromatography.
abstract: Significant progress in the application of viral vectors for gene delivery into mammalian cells and the use of viruses as biopesticides requires downstream processing that can satisfy application-specific demands on performance. In the present work the stability and ion exchange membrane chromatography of a recombinant of Autographa californica M nucleopolyhedrovirus is studied. To adjust the degree of purification the effect of ionic conductivity or pH on the viral infectivity was assessed (0.77-78.00mS/cm, pH 3-8). Infectivity decreased rapidly by several orders of magnitude at below 5mS/cm (i.e., 0.49MPa osmotic pressure change) or at below pH 5.5 (rationalized with particle aggregation). The virus was concentrated and purified via adsorption (0.2-1.1×10(16)pfu/m(3) chromatographic bed volume, 0.6-1.1×10(12)pfu/m(2) membrane area facing the incident fluid flow) and elution at pH 6.1 and 6.35mS/cm from three strong anion exchange membranes. Virus recovery and concentration in accord with the volume reduction were obtained using a polyether sulfone-based membrane with quaternary ammonium ligands. The level of host cell protein (down to below the detection limit) and suspended DNA (below 93pg DNA per 10(6)pfu) are reported for each membrane employed, for the purpose of comparability, under equal adsorption or elution conditions respectively.
authors.affiliation: University of Applied Sciences Mittelhessen, Institute of Bioprocess Engineering and Pharmaceutical Technology, Wiesenstrasse 14, 35390 Giessen, Germany.
authors.forename: Tanja A
authors.initials: TA
authors.lastname: Grein
authors.forename: Ronald
authors.initials: R
authors.lastname: Michalsky
authors.forename: Maria
authors.initials: M
authors.lastname: Vega López
authors.forename: Peter
authors.initials: P
authors.lastname: Czermak
mesh_terms: D000327:Adsorption; D000818:Animals; D002460:Cell Line; D002852:Chromatography, Ion Exchange; D004247:DNA / Q000302:isolation & purification; D004553:Electric Conductivity; D006863:Hydrogen-Ion Concentration; D019476:Insect Proteins / Q000302:isolation & purification; D009036:Moths / Q000821:virology*; D017924:Nucleopolyhedroviruses / Q000235:genetics / Q000302:isolation & purification* / Q000472:pathogenicity; D030781:Organisms, Genetically Modified; D019562:Viral Load
chemical_list: D019476:Insect Proteins; D004247:DNA