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  • Viral Transfer Technology
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Low-speed centrifugation of retroviral vectors absorbed to a particulate substrate: a highly effective means of enhancing retroviral titre

Gene Therapy volume 7, pages 914–923 (2000)Cite this article

Abstract

For many gene therapy applications the effective titre of retroviral vectors is a limiting factor both in vitro and in vivo. Purification and concentration of retrovirus from packaging cell supernatant can overcome this problem. To this end we have investigated a novel procedure which involves complexing retrovirus to a dense and particulate substrate followed by a short low-speed centrifugation. The study reported here uses heat-killed, formaldehyde fixed Staphylococcus aureus (Pansorbin) absorbed to PG13 derived retrovirus. This complex was then used to harvest retrovirus from packaging cell supernatant: centrifugation and washing of this complex allows the retrovirus to be both purified and concentrated. This procedure increases the effective titre of retrovirus by up to 7500-fold after an only 200-fold reduction in volume. The affinity of Pansorbin for retrovirus allows concentration regardless of its encoded genes and makes this protocol applicable to other popular packaging cells and envelope proteins. Possible explanations for the marked increase in titre of concentrated virus and the mechanism governing the complexing of retrovirus to Pansorbin are discussed.

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Acknowledgements

This work was supported by grant 9605 from the Leukaemia Research Fund and The Lewis Family Research Trust. We thank Professor MKL Collins, Department of Immunology, Windeyer Institute of Medical Sciences, University College London for the gift of the FLYA13 and FLYRD18 retroviral packaging cell lines.

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Author notes

  1. M Kuiper

    Present address: Department of Chemical Engineering and Applied Chemistry, Aston University, Aston Triangle, Birmingham University, Birmingham, UK

Authors and Affiliations

  1. Department of Molecular Medicine, Immune Gene Therapy of Cancer Program, King's College School of Medicine and Dentistry, London, UK

    D Darling, C Hughes, J Galea-Lauri, J Gäken, ID Trayner, M Kuiper & F Farzaneh

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Darling, D., Hughes, C., Galea-Lauri, J. et al. Low-speed centrifugation of retroviral vectors absorbed to a particulate substrate: a highly effective means of enhancing retroviral titre. Gene Ther 7, 914–923 (2000). https://doi.org/10.1038/sj.gt.3301201

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