Nuclear pores as versatile reference standards for quantitative superresolution microscopy

• Verfasst von: Thevathasan, Jervis Vermal [VerfasserIn]
• Verfasst von: Ellenberg, Jan [VerfasserIn]
• Verfasst von: Hoess, Philipp [VerfasserIn]
• Verfasst von: Hörner, Sarah J. [VerfasserIn]
• Verfasst von: Wu, Yu-Le [VerfasserIn]
• Titel: Nuclear pores as versatile reference standards for quantitative superresolution microscopy
• Verf.angabe: Jervis Vermal Thevathasan, Maurice Kahnwald, Konstanty Cieśliński, Philipp Hoess, Sudheer Kumar Peneti, Manuel Reitberger, Daniel Heid, Krishna Chaitanya Kasuba, Sarah Janice Hoerner, Yiming Li, Yu-Le Wu, Markus Mund, Ulf Matti, Pedro Matos Pereira, Ricardo Henriques, Bianca Nijmeijer, Moritz Kueblbeck, Vilma Jimenez Sabinina, Jan Ellenberg and Jonas Ries
• E-Jahr: 2019
• Jahr: 27 September 2019
• Umfang: 9 S.
• Fussnoten: Gesehen am 22.04.2020
• Titel Quelle: Enthalten in: Nature methods
• Ort Quelle: London [u.a.] : Nature Publishing Group, 2004
• Jahr Quelle: 2019
• Band/Heft Quelle: 16(2019), 10, Seite 1045-1053
• ISSN Quelle: 1548-7105
• DOI: doi:10.1038/s41592-019-0574-9
• Datenträger: Online-Ressource
• Sprache: eng
• Bibliogr. Hinweis: Korrektur: Thevathasan, Jervis Vermal, 1986 - : Publisher correction
• K10plus-PPN: 1680561774

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.