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. 2017;1575:223-236.
doi: 10.1007/978-1-4939-6857-2_14.

A Biotin Ligase-Based Assay for the Quantification of the Cytosolic Delivery of Therapeutic Proteins

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A Biotin Ligase-Based Assay for the Quantification of the Cytosolic Delivery of Therapeutic Proteins

Wouter P R Verdurmen et al. Methods Mol Biol. .

Abstract

The efficient delivery of external proteins from the external milieu to the cytosol of mammalian cells has great potential for both scientific investigations and future therapies. However, when assessing the cellular uptake of proteins, it is often difficult to distinguish between proteins that are stuck in the endosomes and those that have escaped into the cytosol. Here, we describe a method employing the prokaryotic enzyme biotin ligase that overcomes this problem and which can be employed for a highly sensitive quantification of cytosolic protein delivery.

Keywords: Biotin; Biotin ligase; Cellular internalization; Cytosolic protein delivery; Protein engineering; Streptavidin; Western blot.

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