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. 2022:2510:129-144.
doi: 10.1007/978-1-0716-2384-8_7.

Enhanced Transduction of P2X7-Expressing Cells with Recombinant rAAV Vectors

Anna Marei Mann #  1 , Waldemar Schäfer #  1 , Sahil Adriouch  2 , Kathleen Börner  3   4 , Dirk Grimm  3   4   5 , Ingke Braren  6 , Friedrich Koch-Nolte  7
Affiliations

Enhanced Transduction of P2X7-Expressing Cells with Recombinant rAAV Vectors

Anna Marei Mann et al. Methods Mol Biol. 2022.

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.

Keywords: AAV; Adeno-associated viral vectors; Flow cytometry; Fluorescence microscopy; P2X7; Western blot.

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References

    1. Mingozzi F, High KA (2011) Therapeutic in vivo gene transfer for genetic disease using AAV: Progress and challenges. Nat Rev Genet 12:341–355 - DOI
    1. Kotterman MA, Schaffer DV (2014) Engineering adeno-associated viruses for clinical gene therapy. Nat Rev Genet 15:445–451. https://doi.org/10.1038/nrg3742 - DOI - PubMed - PMC
    1. Colella P, Ronzitti G, Mingozzi F (2018) Emerging issues in AAV-mediated in vivo gene therapy. Mol Ther Meth Clin Dev 8:87–104 - DOI
    1. Herrmann AK, Grimm D (2018) High-throughput dissection of AAV–host interactions: the fast and the curious. J Mol Biol 430:2626–2640 - DOI
    1. Eichhoff AM, Börner K, Albrecht B et al (2019) Nanobody-enhanced targeting of AAV gene therapy vectors. Mol Ther Meth Clin Dev 15:211–220. https://doi.org/10.1016/j.omtm.2019.09.003 - DOI

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