Sulfated membrane adsorbers for economic pseudo‐affinity capture of influenza virus particles

Abstract

Strategies to control outbreaks of influenza, a contagious respiratory tract disease, are focused mainly on prophylactic vaccinations in conjunction with antiviral medications. Currently, several mammalian cell culture-based influenza vaccine production processes are being established, such as the technologies introduced by Novartis Behring (Optaflu) or Baxter International Inc. (Celvapan). Downstream processing of influenza virus vaccines from cell culture supernatant can be performed by adsorbing virions onto sulfated column chromatography beads, such as Cellufine sulfate. This study focused on the development of a sulfated cellulose membrane (SCM) chromatography unit operation to capture cell culture-derived influenza viruses. The advantages of the novel method were demonstrated for the Madin Darby canine kidney (MDCK) cell-derived influenza virus A/Puerto Rico/8/34 (H1N1). Furthermore, the SCM-adsorbers were compared directly to column-based Cellufine sulfate and commercially available cation-exchange membrane adsorbers. Sulfated cellulose membrane adsorbers showed high viral product recoveries. In addition, the SCM-capture step resulted in a higher reduction of dsDNA compared to the tested cation-exchange membrane adsorbers. The productivity of the SCM-based unit operation could be significantly improved by a 30-fold increase in volumetric flow rate during adsorption compared to the bead-based capture method. The higher flow rate even further reduced the level of contaminating dsDNA by about twofold. The reproducibility and general applicability of the developed unit operation were demonstrated for two further MDCK cell-derived influenza virus strains: A/Wisconsin/67/2005 (H3N2) and B/Malaysia/2506/2004. Overall, SCM-adsorbers represent a powerful and economically favorable alternative for influenza virus capture over conventional methods using Cellufine sulfate.

Capturing of cell culture‐derived modified Vaccinia Ankara virus by ion exchange and pseudo‐affinity membrane adsorbers

“…1A) adsorbed to the weak cation exchanger C75-MA and was desorbed by an increased ionic strength via NaCl. A similar result, even more pronounced, was described by Opitz et al (2009) for purification of cell culture-derived influenza virus particles. The overall pI of influenza virus particles as judged by the subtype A 2 /Singapore/57 (5.0; Zhilinskaya et al, 1972) is less acidic than the pI of Vaccinia virus particles.…”

“…Several studies reported antiviral activities of sulfated cellulose and sulfated dextran/dextrines (Baba et al, 1988;Chattopadhyay et al, 2008;Mitsuya et al, 1988;Piret et al, 2000) as well as the binding of virus particles to Cellufine 1 sulfate (O'Neil and Balkovic, 1993;Opitz et al, 2009). Although the precise interaction between these viruses and sulfated cellulose is currently not fully understood, it represents a potential alternative to heparin and heparan sulfate as ligand for MVA-BN 1 virus particles.…”

“…To increase the capacity of the C75-MA and heparin-MA three 75 cm 2 units were serially combined to obtain an overall adsorption area of 225 cm 2 . Viruses have been shown to bind sulfated polysaccharides such as dextran sulfate, heparin, and heparan sulfate (Lycke et al, 1991;Mitsuya et al, 1988;O'Keeffe et al, 1999;O'Neil and Balkovic, 1993;Opitz et al, 2009). Here, a significant difference in the dynamic binding capacities for the tested heparin and sulfated cellulose matrices was observed.…”

“…The housing material was polypropylene. Sulfated cellulose MA (SC-MA) with a diameter of 25 mm (pore size >3 mm, Sartorius Stedim Biotech GmbH) were prepared as described previously (Opitz et al, 2009). The adsorption area was 75 cm 2 and 15 membranes were stacked in a stainless steel membrane holder (Cat.# 1980-002, GE Healthcare, Munich, Germany).…”

“…One possibility to economize or even to eliminate the nuclease treatment in a new production process might be the application of adsorption chromatography methods, ideally using chromatography media with predominant convective material transport like membrane adsorbers (MA). There are several examples for the application of ion exchange and affinity MA for the purification of virus particles like adenoviral vectors (Peixoto et al, 2008;Sellick, 2006), Aedes aegypti densonucleosis virus (Enden et al, 2005), baculovirus (Wu et al, 2007), and influenza virus (Kalbfuss et al, 2007a;Opitz et al, 2007aOpitz et al, , 2009. With regard to the low isoelectric point (pI) of $4.0, as estimated for the Lister strain of Vaccinia virus (Douglas et al, 1969), anion exchange MA might be a promising option.…”