Comparative Assessment of Fluorescent Transgene Methods for Quantitative Imaging in Human Cells

Overview

Journal Mol Biol Cell

Specialties Cell Biology
Molecular Biology

Date 2014 Sep 19

PMID 25232003

Citations 24

Authors

Robert Mahen

Birgit Koch

Malte Wachsmuth

Antonio Z Politi

Alexis Perez-Gonzalez

Julia Mergenthaler

Yin Cai

Jan Ellenberg

Affiliations

Soon will be listed here.

Abstract

Fluorescence tagging of proteins is a widely used tool to study protein function and dynamics in live cells. However, the extent to which different mammalian transgene methods faithfully report on the properties of endogenous proteins has not been studied comparatively. Here we use quantitative live-cell imaging and single-molecule spectroscopy to analyze how different transgene systems affect imaging of the functional properties of the mitotic kinase Aurora B. We show that the transgene method fundamentally influences level and variability of expression and can severely compromise the ability to report on endogenous binding and localization parameters, providing a guide for quantitative imaging studies in mammalian cells.

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