tetano
Editor, Senior Moderator
J Biotechnol. 2015 Dec 19. pii: S0168-1656(15)30221-2. doi: 10.1016/j.jbiotec.2015.12.022. [Epub ahead of print]
[h=1]A membrane-based purification process for cell culture-derived influenza A virus.[/h] Weigel T[SUP]1[/SUP], Solomaier T[SUP]2[/SUP], Wehmeyer S[SUP]3[/SUP], Peuker A[SUP]4[/SUP], Wolff MW[SUP]5[/SUP], Reichl U[SUP]5[/SUP].
[h=3]Author information[/h]
[h=3]Abstract[/h] A simple membrane-based purification process for cell culture-derived influenza virus was established that relies on only two chromatographic unit operations to achieve the contamination limits required according to regulatory authorities. After clarification and concentration, a pseudo-affinity membrane adsorber (sulfated cellulose, SCMA) was applied for virus capture. The subsequent polishing step consisted of a salt-tolerant anion exchange membrane adsorber (STMA) to bind residual DNA. For the presented process neither a buffer exchange step nor a nuclease step for further DNA digestion were required. As a starting point, a two-salt strategy (including a polyvalent ion) was employed to screen STMA conditions in a 96-well plate format. After optimization on chromatographic laboratory scale, the virus recovery was up to 97% with a residual DNA level below 0.82%. In addition, the STMA was characterized regarding its dynamic binding capacity and the impact of flow rate on yields and contamination levels. Overall, the total virus yield for influenza virus A/PR/8/34 (H1/N1) of this two-step membrane process was 75%, while the protein and the DNA contamination level could be reduced to 24% and at least 0.5%, respectively. With 19.8μg protein and 1.2ng DNA per monovalent dose, this purity level complies with the limits of the European Pharmacopeia for cell culture-derived vaccines for human use. Overall, the presented downstream process might serve as a generic and economic platform technology for production of cell culture-derived viruses and viral vectors.
Copyright ? 2015. Published by Elsevier B.V.
[h=4]KEYWORDS:[/h] Downstream process; Influenza virus vaccine purification process; Primary amine anion exchange membrane; Pseudo-affinity membrane chromatography; Salt-tolerant membrane chromatography; Sulfated cellulose membrane adsorber
PMID: 26712479 [PubMed - as supplied by publisher]
[h=1]A membrane-based purification process for cell culture-derived influenza A virus.[/h] Weigel T[SUP]1[/SUP], Solomaier T[SUP]2[/SUP], Wehmeyer S[SUP]3[/SUP], Peuker A[SUP]4[/SUP], Wolff MW[SUP]5[/SUP], Reichl U[SUP]5[/SUP].
[h=3]Author information[/h]
[h=3]Abstract[/h] A simple membrane-based purification process for cell culture-derived influenza virus was established that relies on only two chromatographic unit operations to achieve the contamination limits required according to regulatory authorities. After clarification and concentration, a pseudo-affinity membrane adsorber (sulfated cellulose, SCMA) was applied for virus capture. The subsequent polishing step consisted of a salt-tolerant anion exchange membrane adsorber (STMA) to bind residual DNA. For the presented process neither a buffer exchange step nor a nuclease step for further DNA digestion were required. As a starting point, a two-salt strategy (including a polyvalent ion) was employed to screen STMA conditions in a 96-well plate format. After optimization on chromatographic laboratory scale, the virus recovery was up to 97% with a residual DNA level below 0.82%. In addition, the STMA was characterized regarding its dynamic binding capacity and the impact of flow rate on yields and contamination levels. Overall, the total virus yield for influenza virus A/PR/8/34 (H1/N1) of this two-step membrane process was 75%, while the protein and the DNA contamination level could be reduced to 24% and at least 0.5%, respectively. With 19.8μg protein and 1.2ng DNA per monovalent dose, this purity level complies with the limits of the European Pharmacopeia for cell culture-derived vaccines for human use. Overall, the presented downstream process might serve as a generic and economic platform technology for production of cell culture-derived viruses and viral vectors.
Copyright ? 2015. Published by Elsevier B.V.
[h=4]KEYWORDS:[/h] Downstream process; Influenza virus vaccine purification process; Primary amine anion exchange membrane; Pseudo-affinity membrane chromatography; Salt-tolerant membrane chromatography; Sulfated cellulose membrane adsorber
PMID: 26712479 [PubMed - as supplied by publisher]