DOI: 10.5281/zenodo.5068524
title: Sharpening emitter localization in front of a tuned mirror - NPC dataset

description: This is a depository for two single molecule localisation microscopy datasets of nuclear pore complex (NPC) structures for single particle averaging. The data was published in: Heil, H.S., Schreiber, B., Götz, R. <em>et al.</em> Sharpening emitter localisation in front of a tuned mirror. <em>Light Sci Appl</em> <strong>7, </strong>99 (2018). https://doi.org/10.1038/s41377-018-0104-z Both datasets have two different levels of localisation precision as one is a conventional STORM experiment and the second a mirror-enhanced STORM experiment. A detailed description of the sample preparation and imaging conditions can be found in the related publication. In short the NPC structures are placed on the surface of a glas coverslip or nano-mirror coated coverslip by manual isolation and spreading of nuclear envelopes from xenopus laevis oocytes, fixed and stained by indirect immunolabeling. The primary antibody targets GP210, the secondary F(ab')<sub>2</sub> fragment is conjugated with Alexa Fluor 647. In this depository I'm providing the raw images data, localisation data and super-resolved reconstruction for the two experiments, as well as the localisation data and super-resolved reconstruction of single NPC rings. I'm also providing a MatLab script that allows to select single NPC positions in the super-resolved image and export the localization data of the single NPC ROI: <strong>P01_ImageAlignment_PickElements.m</strong> Information about the dataset is also available here: <strong>NPC Image Alignment Dataset_Info.pdf.</strong> Image parameters: 102 nm pixel size, EM Gain 100, Photoelectrons per A/D count: 15.01 Column structure of the localisation text files: Id,Frame, x [nm], y [nm], sigma [nm], intensity [photon], offset [photon], bkgstd [photon], chi2, Uncertainty [nm], detections Files: <strong>NPCData_glass_EPI.tif</strong> -&gt; NPC on glass coverslip, low power EPI illumination, widefield image, 20 ms exposure <strong>NPCData_glass_STORM.tif</strong> -&gt; NPC on glass coverslip, high power EPI illumination, 5 ms exposure, 20000 frames <strong>NPCData_glass_STORM_loc.csv</strong> -&gt; ThunderSTORM Localisation data of NPCData_glass_STORM.tif, parameters specified NPCData_glass_STORM_loc-protocol.txt <strong>NPCData_glass_STORM_20xNormalizedGaussian.tif</strong> -&gt; 20x Nomalized Gaussian reconstruction of localization data from NPCData_glass_STORM.tif (ThunderSTORM), pixelsize 5.1 nm <strong>NPCData_glass_STORM_singleRings.zip</strong> -&gt; Localisation data and 20x 20x Nomalized Gaussian reconstruction of single NPC ROIs picked out of the NPCData_glass_STORM dataset, ROI size is 240*240 nm<sup>2</sup> <strong>NPCData_nanomirror_EPI.tif</strong> -&gt; NPC on nanomirror coated coverslip, low power EPI illumination, widefield image, 20 ms exposure <strong>NPCData_nanomirror_STORM.tif</strong> -&gt; NPC on nanomirror coated coverslip, high power EPI illumination, 5 ms exposure, 20000 frames <strong>NPCData_nanomirror_STORM_loc.csv</strong> -&gt; ThunderSTORM Localisation data of NPCData_nanomirror_STORM.tif, parameters specified NPCData_nanomirror_STORM_loc-protocol.txt <strong>NPCData_nanomirror_STORM_20xNormalizedGaussian.tif</strong> -&gt; 20x Nomalized Gaussian reconstruction of localisation data from NPCData_nanomirror_STORM.tif (ThunderSTORM), pixelsize 5.1 nm <strong>NPCData_nanomirror_STORM_singleRings.zip</strong> -&gt; Localisation data and 20x 20x Nomalized Gaussian reconstruction of single NPC ROIs picked out of the NPCData_nanomirror_STORM dataset, ROI size is 240*240 nm<sup>2</sup>