An efficient process for the purification of helper-dependent adenoviral vector and removal of helper virus by iodixanol ultracentrifugation

Journal of Virological Methods
E DormondAmine Kamen

Abstract

The preparation of large amount of purified helper-dependent adenoviral vector material is hampered by the lack of development of downstream processes with proven records on separation and recovery efficiencies. In order to facilitate the use of clinical-grade helper-dependent virus material for large-scale in vivo studies, a three-step purification scheme consisting of (1) an anion-exchange chromatography for initial capturing of virus, (2) a shallow iodixanol density gradient ultracentrifugation for the removal of helper virus from helper-dependent virus, and (3) a size-exclusion chromatography for the removal of iodixanol and residual protein contaminants as a polishing step was developed. The use of a fast iodixanol density ultracentrifugation step was highly effective in separating infectious helper-dependent virus from contaminating helper virus. The overall downstream processing scheme gave 80% infectious particle yield. The contamination ratio of helper virus in the helper-dependent virus preparation are reduced from 2.57 to 0.03% corresponding to a reduction of helper virus by factors of 85 by two iodixanol purification steps. It was also demonstrated that size-exclusion chromatography is an excellent step for the remo...Continue Reading

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Citations

Apr 29, 2011·Gene Therapy·M Gonzalez-AparicioR Hernandez-Alcoceba
Nov 16, 2010·Expert Opinion on Drug Delivery·Mohube Betty MowaPatrick Arbuthnot
Feb 7, 2013·Journal of Fish Diseases·J J Dantas-LimaH J Nauwynck
Mar 10, 2015·Journal of Biotechnology·Piergiuseppe NestolaJosé P B Mota
Jul 8, 2016·Proceedings of the National Academy of Sciences of the United States of America·Ori BratenAaron Ciechanover
May 13, 2014·Journal of Chromatography. a·Piergiuseppe NestolaJosé P B Mota
Sep 25, 2017·Biotechnology and Bioengineering·David Sharon, Amine Kamen
May 28, 2020·International Journal of Molecular Sciences·Ana RicobarazaRuben Hernandez-Alcoceba
May 5, 2020·Journal of Extracellular Vesicles·Heikki SaariMarjo Yliperttula

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