Abstract
Adeno-associated viruses (AAV) are useful vectors for transducing cells in vitro and in vivo. Targeting of specific cell subsets with AAV is limited by the broad tropism of AAV serotypes. Nanobodies are single immunoglobulin variable domains from heavy chain antibodies that naturally occur in camelids. Their small size and high solubility allow easy reformatting into fusion proteins. In this chapter we provide protocols for inserting a P2X7-specific nanobody into a surface loop of the VP1 capsid protein of AAV2. Such nanobody-displaying recombinant AAV allow 50- to 500-fold stronger transduction of P2X7-expressing cells than the parental AAV. We provide protocols for monitoring the transduction of P2X7-expressing cells by nanobody-displaying rAAV by flow cytometry and fluorescence microscopy.
Anna Marei Mann is the same first author as A.M.E. in Eichhoff et al. Molecular Therapy: Methods & Clinical Development Vol. 15 December 2019.
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Acknowledgments
Supported by a stipend from the Claussen-Simon-Stiftung to A.M.M., and grants from the Deutsche Forschungsgemeinschaft to F.K.-N. (No310/13-1, SFB1328Z02), from the German Center for Infection Research to K.B. and D.G. (TTU-HIV 04.803 and TTU-HIV 04.815), and from the Agence Nationale de la Recherche to S.A. (ANR-18-CE92-0046).
We thank Fabienne Seyfried and Josi Gebhard (Institute of Immunology, UKE, Hamburg) for excellent technical assistance.
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Mann, A.M. et al. (2022). Enhanced Transduction of P2X7-Expressing Cells with Recombinant rAAV Vectors. In: Nicke, A. (eds) The P2X7 Receptor. Methods in Molecular Biology, vol 2510. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-2384-8_7
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DOI: https://doi.org/10.1007/978-1-0716-2384-8_7
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