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A membrane-based purification process for cell culture-derived influenza A virus

Paper ID Volume ID Publish Year Pages File Format Full-Text
22744 43380 2016 9 PDF Available
Title
A membrane-based purification process for cell culture-derived influenza A virus
Abstract

•Two-step membrane-based chromatographic purification process for influenza virus.•Separation of virus particles and DNA by a novel salt-tolerant membrane adsorber.•High virus yield (75%) and reduction of DNA content to at least 0.5%.•Compliance with European pharmacopeia requirements for DNA and total protein.

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 19.8 μg protein and 1.2 ng 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.

Keywords
LOQ, limit of quantification; SCMA, sulfated cellulose membrane adsorber; STMA, salt-tolerant membrane adsorber; DBC, dynamic binding capacity; CV, column volumes; UF, ultrafiltration; A/PR, influenza virus A/PR/8/34; GMEM, glasgow minimum essential mediu
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A membrane-based purification process for cell culture-derived influenza A virus
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Publisher
Database: Elsevier - ScienceDirect
Journal: Journal of Biotechnology - Volume 220, 20 February 2016, Pages 12–20
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering
Get Full-Text Now
Don't Miss Today's Special Offer
Price was $35.95
You save - $31
Price after discount Only $4.95
100% Money Back Guarantee
Full-text PDF Download
Online Support
Any Questions? feel free to contact us