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Methods Mol Biol. 2009;485:151-9. doi: 10.1007/978-1-59745-170-3_11.

Bisarsenical labeling of HIV-1 for real-time fluorescence microscopy.

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  • 1Virology Department, Institut Pasteur, Molecular Virology and Vectorology Group, Paris, France.

Abstract

Imaging studies have benefited from the development of a novel technique for non-destructive labeling of proteins within living cells, based on the use of a reagent called FlAsH-EDT2, a bisarsenical derivative of fluorescein capable of binding with high affinity and specificity to a tetracysteine motif in the protein of interest. This technique has been adapted for the stable, sensitive and specific molecular tagging of HIV-1 IN enabling the tracking of incoming viral particles inside infected living cells. Here we present the experimental steps required for the efficient labeling of HIV-1 IN, namely, molecular insertion of a tetracysteine tag, production of viruses, labeling in vitro of tagged viruses, infection of target cells and visualization of particles by fluorescence microscopy.

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