title: Porcine parvovirus flocculation and removal in the presence of osmolytes.
abstract: Viruses can be modified into viral vaccines or gene therapy vectors in order to treat acquired or genetic diseases. To satisfy the current market demand, an improvement in current vaccine manufacturing is needed. Chromatography and nanofiltration are not suitable for all types of viruses. In this study, we propose to use virus flocculation with osmolytes, followed by microfiltration, as a potential virus purification process. We hypothesize that osmolytes strongly bind to water, thus leading to the formation of a hydration layer around the virus particles and stimulation of aggregation. We have discovered that osmolytes, including sugars, sugar alcohols and amino acids, preferentially flocculate porcine parvovirus (PPV), and demonstrate a >80% removal with a 0.2 μm filter while leaving model proteins in solution. This large pore size filter increases the flux and decreases the transmembrane pressure of typical virus filters. The best flocculants were tested for their ability to aggregate PPV at different concentrations, shear stress, pH and ionic strength. We were able to remove 96% of PPV in 3.0M glycine at a pH of 5. Glycine is also an excipient, and therefore may not require removal later in the process. Virus flocculation using osmolytes, followed by microfiltration could be used as an integrated process for virus purification.
authors.affiliation: Department of Chemical Engineering, Michigan Technological University, 1400 Townsend Dr. Houghton, MI 49931, USA.
authors.forename: Maria F
authors.initials: MF
authors.lastname: Gencoglu
authors.affiliation: Department of Chemical Engineering, Michigan Technological University, 1400 Townsend Dr. Houghton, MI 49931, USA.
authors.forename: Eric
authors.initials: E
authors.lastname: Pearson
authors.affiliation: Department of Chemical Engineering, Michigan Technological University, 1400 Townsend Dr. Houghton, MI 49931, USA. Electronic address: heldt@mtu.edu.
authors.forename: Caryn L
authors.initials: CL
authors.lastname: Heldt
mesh_terms: D000409:Alanine / Q000378:metabolism; D000818:Animals; D002460:Cell Line; D005427:Flocculation; D005998:Glycine / Q000378:metabolism; D057166:High-Throughput Screening Assays; D006863:Hydrogen-Ion Concentration; D008353:Mannitol / Q000378:metabolism; D009994:Osmolar Concentration; D028323:Parvovirus, Porcine / Q000254:growth & development* / Q000302:isolation & purification* / Q000378:metabolism; D016062:Porosity; D012965:Sodium Chloride / Q000737:chemistry; D013552:Swine; D014776:Virus Cultivation / Q000379:methods*
chemical_list: D008353:Mannitol; D012965:Sodium Chloride; D000409:Alanine; D005998:Glycine
keywords: Aggregation; Alanine; High throughput; Mannitol; PPV; Virus precipitation