Nuclear Pores As Versatile Reference Standards for Quantitative Superresolution Microscopy

Overview

Journal Nat Methods

Specialties Biomedical Engineering
Pathology

Date 2019 Sep 29

PMID 31562488

Citations 133

Authors

Jervis Vermal Thevathasan

Maurice Kahnwald

Konstanty Cieslinski

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

Jonas Ries

Affiliations

Soon will be listed here.

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.

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