scholarly journals Fast Analysis of 2D and 3D Single-Molecule Localization Microscopy Data with Huygens Localizer

2020 ◽  
Vol 28 (2) ◽  
pp. 20-25
Author(s):  
Peter J. Verveer ◽  
Daniel Sevilla-Sánchez ◽  
Paul C. Bloembergen ◽  
Nancy Adam ◽  
Vincent T.G. Schoonderwoert ◽  
...  
Keyword(s):  
Single Molecule ◽  
Fast Analysis ◽  
Localization Microscopy ◽  
Microscopy Data ◽  
Image Position ◽  
2D And 3D ◽  
Single Molecule Localization Microscopy

Abstract

scholarly journals StormGraph: A graph-based algorithm for quantitative clustering analysis of diverse single-molecule localization microscopy data

2019 ◽  
Author(s):  
Joshua M. Scurll ◽  
Libin Abraham ◽  
Da Wei Zheng ◽  
Reza Tafteh ◽  
Keng C. Chou ◽  
...  
Keyword(s):  
Single Molecule ◽  
Input Parameter ◽  
Surface Protein ◽  
Ground Truth ◽  
Antigen Receptors ◽  
Localization Microscopy ◽  
3D Data ◽  
Microscopy Data ◽  
2D And 3D ◽  
Single Molecule Localization Microscopy

AbstractClustering of proteins is crucial for many cellular processes and can be imaged at nanoscale resolution using single-molecule localization microscopy (SMLM). Ideally, molecular clustering in regions of interest (ROIs) from SMLM images would be assessed using computational methods that are robust to sample and experimental heterogeneity, account for uncertainties in localization data, can analyze both 2D and 3D data, and have practical computational requirements in terms of time and hardware. While analyzing surface protein clustering on B lymphocytes using SMLM, we encountered limitations with existing cluster analysis methods. This inspired us to develop StormGraph, an algorithm using graph theory and community detection to identify clusters in heterogeneous sets of 2D and 3D SMLM data while accounting for localization uncertainties. StormGraph generates both multi-level and single-level clusterings and can quantify cluster overlap for two-color SMLM data. Importantly, StormGraph automatically determines scale-dependent thresholds from the data using scale-independent input parameters. This makes identical choices of input parameter values suitable for disparate ROIs, eliminating the need to tune parameters for different ROIs in heterogeneous SMLM datasets. We show that StormGraph outperforms existing algorithms at analyzing heterogeneous sets of simulated SMLM ROIs where ground-truth clusters are known. Applying StormGraph to real SMLM data in 2D, we reveal that B-cell antigen receptors (BCRs) reside in a heterogeneous combination of small and large clusters following stimulation, which suggests for the first time that two conflicting models of BCR activation are not mutually exclusive. We also demonstrate application of StormGraph to real two-color and 3D SMLM data.

scholarly journals Super-resolution fight club: assessment of 2D and 3D single-molecule localization microscopy software

2019 ◽  
Vol 16 (5) ◽  
pp. 387-395 ◽  
Author(s):  
Daniel Sage ◽  
Thanh-An Pham ◽  
Hazen Babcock ◽  
Tomas Lukes ◽  
Thomas Pengo ◽  
...  
Keyword(s):  
Single Molecule ◽  
Super Resolution ◽  
Localization Microscopy ◽  
Fight Club ◽  
2D And 3D ◽  
Single Molecule Localization Microscopy

scholarly journals SMoLR: visualization and analysis of single-molecule localization microscopy data in R

2019 ◽  
Vol 20 (1) ◽  
Author(s):  
Maarten W. Paul ◽  
H. Martijn de Gruiter ◽  
Zhanmin Lin ◽  
Willy M. Baarends ◽  
Wiggert A. van Cappellen ◽  
...  
Keyword(s):  
Single Molecule ◽  
Localization Microscopy ◽  
Microscopy Data ◽  
Single Molecule Localization Microscopy

scholarly journals Super-resolution fight club: A broad assessment of 2D & 3D single-molecule localization microscopy software

2018 ◽  
Author(s):  
Daniel Sage ◽  
Thanh-An Pham ◽  
Hazen Babcock ◽  
Tomas Lukes ◽  
Thomas Pengo ◽  
...  
Keyword(s):  
Single Molecule ◽  
Double Helix ◽  
Super Resolution ◽  
Large Set ◽  
Holistic View ◽  
Localization Microscopy ◽  
Fight Club ◽  
Community Effort ◽  
2D And 3D ◽  
Single Molecule Localization Microscopy

ABSTRACTWith the widespread uptake of 2D and 3D single molecule localization microscopy, a large set of different data analysis packages have been developed to generate super-resolution images. To guide researchers on the optimal analytical software for their experiments, we have designed, in a large community effort, a competition to extensively characterise and rank these options. We generated realistic simulated datasets for popular imaging modalities – 2D, astigmatic 3D, biplane 3D, and double helix 3D – and evaluated 36 participant packages against these data. This provides the first broad assessment of 3D single molecule localization microscopy software, provides a holistic view of how the latest 2D and 3D single molecule localization software perform in realistic conditions, and ultimately provides insight into the current limits of the field.

scholarly journals A workflow for sizing oligomeric biomolecules based on cryo single molecule localization microscopy

2020 ◽  
Author(s):  
Magdalena C. Schneider ◽  
Roger Telschow ◽  
Gwenael Mercier ◽  
Montserrat López-Martinez ◽  
Otmar Scherzer ◽  
...  
Keyword(s):  
Single Molecule ◽  
High Reliability ◽  
Limiting Factor ◽  
Dye Molecules ◽  
New Approach ◽  
Localization Microscopy ◽  
Additional Information ◽  
Localization Precision ◽  
Microscopy Data ◽  
Single Molecule Localization Microscopy

ABSTRACTSingle molecule localization microscopy (SMLM) has enormous potential for resolving subcellular structures below the diffraction limit of light microscopy: Localization precision in the low digit nanometer regime has been shown to be achievable. In order to record localization microscopy data, however, sample fixation is inevitable to prevent molecular motion during the rather long recording times of minutes up to hours. Eventually, it turns out that preservation of the sample’s ultrastructure during fixation becomes the limiting factor. We propose here a workflow for data analysis, which is based on SMLM performed at cryogenic temperatures. Since molecular dipoles of the fluorophores are fixed at low temperatures, such an approach offers the possibility to use the orientation of the dipole as an additional information for image analysis. In particular, assignment of localizations to individual dye molecules becomes possible with high reliability. We quantitatively characterized the new approach based on the analysis of simulated oligomeric structures. Side lengths can be determined with a relative error of less than 1% for tetramers with a nominal side length of 5 nm, even if the assumed localization precision for single molecules is more than 2 nm.

scholarly journals Quantitative fibre analysis of single-molecule localization microscopy data

2018 ◽  
Vol 8 (1) ◽  
Author(s):  
Ruby Peters ◽  
Juliette Griffié ◽  
Garth L. Burn ◽  
David J. Williamson ◽  
Dylan M. Owen
Keyword(s):  
Single Molecule ◽  
Localization Microscopy ◽  
Microscopy Data ◽  
Fibre Analysis ◽  
Single Molecule Localization Microscopy

Coordinate-based colocalization analysis of single-molecule localization microscopy data

2011 ◽  
Vol 137 (1) ◽  
pp. 1-10 ◽  
Author(s):  
Sebastian Malkusch ◽  
Ulrike Endesfelder ◽  
Justine Mondry ◽  
Márton Gelléri ◽  
Peter J. Verveer ◽  
...  
Keyword(s):  
Single Molecule ◽  
Localization Microscopy ◽  
Microscopy Data ◽  
Single Molecule Localization Microscopy
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