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Issue 5, 2015
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In-gel detection of biotin–protein conjugates with a green fluorescent streptavidin probe

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Abstract

Exploitation of the (strept)avidin–biotin interaction is extremely valuable in a variety of biotechnological applications. Biotin is often covalently linked to proteins or nucleic acids. Determination of the degree of biotinylation of such macromolecules is essential for downstream applications. There is currently a gap in simple yet efficient assays for rapidly quantitating protein biotinylation, as staple methods may produce unclear results or rely on immuno- or competitive assays. We present a simple and reliable electrophoretic method to determine the relative extent of biotinylation of macromolecules. The method relies on complex formation between a biotinylated macromolecule and a streptavidin probe resulting in an electrophoretic mobility shift of the complex detectable by SDS-PAGE. Finally, a green fluorescent protein labelled streptavidin probe was developed to eliminate the need for staining and reduce assay time.

Graphical abstract: In-gel detection of biotin–protein conjugates with a green fluorescent streptavidin probe

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Supplementary files

Article information


Submitted
10 Nov 2014
Accepted
28 Jan 2015
First published
28 Jan 2015

This article is Open Access

Anal. Methods, 2015,7, 2087-2092
Article type
Paper
Author version available

In-gel detection of biotin–protein conjugates with a green fluorescent streptavidin probe

A. E. Sorenson, S. P. Askin and P. M. Schaeffer, Anal. Methods, 2015, 7, 2087
DOI: 10.1039/C4AY02666G

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