Thevathasan, JV;
Kahnwald, M;
Cieslinski, K;
Hoess, P;
Peneti, SK;
Reitberger, M;
Heid, D;
... Ries, J; + view all
(2019)
Nuclear pores as versatile reference standards for quantitative superresolution microscopy.
Nature Methods
, 16
pp. 1045-1053.
10.1038/s41592-019-0574-9.
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Abstract
Quantitative fluorescence and superresolution microscopy are often limited by insufficient data quality or artifacts. In this context, it is essential to have biologically relevant control samples to benchmark and optimize the quality of microscopes, labels and imaging conditions. Here, we exploit the stereotypic arrangement of proteins in the nuclear pore complex as in situ reference structures to characterize the performance of a variety of microscopy modalities. We created four genome edited cell lines in which we endogenously labeled the nucleoporin Nup96 with mEGFP, SNAP-tag, HaloTag or the photoconvertible fluorescent protein mMaple. We demonstrate their use (1) as three-dimensional resolution standards for calibration and quality control, (2) to quantify absolute labeling efficiencies and (3) as precise reference standards for molecular counting. These cell lines will enable the broader community to assess the quality of their microscopes and labels, and to perform quantitative, absolute measurements.
| Type: | Article |
|---|---|
| Title: | Nuclear pores as versatile reference standards for quantitative superresolution microscopy |
| Open access status: | An open access version is available from UCL Discovery |
| DOI: | 10.1038/s41592-019-0574-9 |
| Publisher version: | https://doi.org/10.1038/s41592-019-0574-9 |
| Language: | English |
| Additional information: | This version is the author accepted manuscript. For information on re-use, please refer to the publisher’s terms and conditions. |
| UCL classification: | UCL UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Life Sciences UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Life Sciences > Lab for Molecular Cell Bio MRC-UCL |
| URI: | https://discovery.ucl.ac.uk/id/eprint/10084914 |
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