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Large-scale production of lentiviral vectors using multilayer cell factories
Lentiviral-mediated gene therapy has been proposed for the treatment of a range of diseases, and due to its genome integration properties, it offers the potential for long-lasting benefit from a once-off treatment. Production methods for pre-clinical studies in animal models, and ultimately for huma...
Autores principales: | , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Journal of Biological Methods
2018
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6706103/ https://www.ncbi.nlm.nih.gov/pubmed/31453241 http://dx.doi.org/10.14440/jbm.2018.236 |
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author | Rout-Pitt, Nathan McCarron, Alexandra McIntyre, Chantelle Parsons, David Donnelley, Martin |
author_facet | Rout-Pitt, Nathan McCarron, Alexandra McIntyre, Chantelle Parsons, David Donnelley, Martin |
author_sort | Rout-Pitt, Nathan |
collection | PubMed |
description | Lentiviral-mediated gene therapy has been proposed for the treatment of a range of diseases, and due to its genome integration properties, it offers the potential for long-lasting benefit from a once-off treatment. Production methods for pre-clinical studies in animal models, and ultimately for human clinical trials, must be capable of producing large quantities of high-quality lentiviral vector in an efficient and cost-effective manner. We report here a medium-scale method (from 1.5 L to 6 L of vector supernatant) for lentiviral vector production in adherent cell cultures using the NUNC™ EasyFill™ Cell Factory™ from Thermo Fisher Scientific. Downstream purification uses a Mustang Q XT5 anion exchange capsule from Pall, and an ultracentrifugation step to concentrate the vector. This method is capable of producing lentiviral vector with concentrated titres of 10(8)–10(9) TU/ml, with reduced manual handling compared to single monolayer flask methods. |
format | Online Article Text |
id | pubmed-6706103 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2018 |
publisher | Journal of Biological Methods |
record_format | MEDLINE/PubMed |
spelling | pubmed-67061032019-08-26 Large-scale production of lentiviral vectors using multilayer cell factories Rout-Pitt, Nathan McCarron, Alexandra McIntyre, Chantelle Parsons, David Donnelley, Martin J Biol Methods Protocol Lentiviral-mediated gene therapy has been proposed for the treatment of a range of diseases, and due to its genome integration properties, it offers the potential for long-lasting benefit from a once-off treatment. Production methods for pre-clinical studies in animal models, and ultimately for human clinical trials, must be capable of producing large quantities of high-quality lentiviral vector in an efficient and cost-effective manner. We report here a medium-scale method (from 1.5 L to 6 L of vector supernatant) for lentiviral vector production in adherent cell cultures using the NUNC™ EasyFill™ Cell Factory™ from Thermo Fisher Scientific. Downstream purification uses a Mustang Q XT5 anion exchange capsule from Pall, and an ultracentrifugation step to concentrate the vector. This method is capable of producing lentiviral vector with concentrated titres of 10(8)–10(9) TU/ml, with reduced manual handling compared to single monolayer flask methods. Journal of Biological Methods 2018-04-10 /pmc/articles/PMC6706103/ /pubmed/31453241 http://dx.doi.org/10.14440/jbm.2018.236 Text en © 2018 The Journal of Biological Methods, All rights reserved. https://creativecommons.org/licenses/by/3.0/ This work is licensed under a Creative Commons Attribution 3.0 License. |
spellingShingle | Protocol Rout-Pitt, Nathan McCarron, Alexandra McIntyre, Chantelle Parsons, David Donnelley, Martin Large-scale production of lentiviral vectors using multilayer cell factories |
title | Large-scale production of lentiviral vectors using multilayer cell factories |
title_full | Large-scale production of lentiviral vectors using multilayer cell factories |
title_fullStr | Large-scale production of lentiviral vectors using multilayer cell factories |
title_full_unstemmed | Large-scale production of lentiviral vectors using multilayer cell factories |
title_short | Large-scale production of lentiviral vectors using multilayer cell factories |
title_sort | large-scale production of lentiviral vectors using multilayer cell factories |
topic | Protocol |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6706103/ https://www.ncbi.nlm.nih.gov/pubmed/31453241 http://dx.doi.org/10.14440/jbm.2018.236 |
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