Abstract
Ectopically expressed intracellular recombinant antibodies, or intrabodies, are powerful tools to visualize proteins and study their function in fixed or living cells. However, many intrabodies are insoluble and aggregate in the reducing environment of the cytosol. To solve this problem, we describe an approach based on GFP-tagged intrabodies. In this protocol, the GFP is used both as a folding-reporter to select correctly folded intrabodies and as a fluorescent tag to localize the scFv and its associated antigen in eukaryotic cells. Starting from a scFv gene cloned in a retroviral vector, we describe retrovirus production, cell line transduction, and soluble intrabody characterization by microscopy and FACS analysis.
The original version of this chapter was revised. The erratum to this chapter is available at: DOI 10.1007/978-1-4939-6857-2_27
An erratum to this chapter can be found at http://dx.doi.org/10.1007/978-1-4939-6857-2_27
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Renaud, E., Martineau, P., Guglielmi, L. (2017). Solubility Characterization and Imaging of Intrabodies Using GFP-Fusions. In: Tiller, T. (eds) Synthetic Antibodies. Methods in Molecular Biology, vol 1575. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-6857-2_9
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DOI: https://doi.org/10.1007/978-1-4939-6857-2_9
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