Robust online image processing for high-throughput super-resolution localization microscopy

2019 ◽  
Author(s):  
Hongqiang Ma ◽  
Jianquan Xu ◽  
Yang Liu
Keyword(s):  
Image Processing ◽  
High Throughput ◽  
Super Resolution ◽  
Localization Microscopy ◽  
Online Image Processing

scholarly journals WindSTORM: Robust online image processing for high-throughput nanoscopy

2019 ◽  
Vol 5 (4) ◽  
pp. eaaw0683 ◽  
Author(s):  
Hongqiang Ma ◽  
Jianquan Xu ◽  
Yang Liu
Keyword(s):  
Image Processing ◽  
High Throughput ◽  
High Speed ◽  
High Density ◽  
Frame Rate ◽  
Large Field ◽  
Localization Accuracy ◽  
Image Characteristics ◽  
Emitter Localization ◽  
Online Image Processing

High-throughput nanoscopy becomes increasingly important for unraveling complex biological processes from a large heterogeneous cell population at a nanoscale resolution. High-density emitter localization combined with a large field of view and fast imaging frame rate is commonly used to achieve a high imaging throughput, but the image processing speed and the presence of heterogeneous background in the dense emitter scenario remain a bottleneck. Here, we present a simple non-iterative approach, referred to as WindSTORM, to achieve high-speed high-density emitter localization with robust performance for various image characteristics. We demonstrate that WindSTORM improves the computation speed by two orders of magnitude on CPU and three orders of magnitude upon GPU acceleration to realize online image processing, without compromising localization accuracy. Further, WindSTORM is highly robust to maximize the localization accuracy and minimize the image artifacts in the presence of nonuniform background. WindSTORM paves the way for next generation high-throughput nanoscopy.

scholarly journals WindSTORM: Robust online image processing for high-throughput nanoscopy

2018 ◽  
Author(s):  
Hongqiang Ma ◽  
Jianquan Xu ◽  
Yang Liu
Keyword(s):  
Image Processing ◽  
High Throughput ◽  
High Speed ◽  
High Density ◽  
Frame Rate ◽  
Large Field ◽  
Localization Accuracy ◽  
Uniform Background ◽  
Emitter Localization ◽  
Online Image Processing

AbstractHigh-throughput nanoscopy becomes increasingly important for unraveling complex biological processes from a large heterogeneous cell population at a nanoscale resolution. High-density emitter localization combined with a large field of view and fast imaging frame rate is commonly used to achieve a high imaging throughput, but the image processing speed in the dense emitter scenario remains a bottleneck. Here we present a simple non-iterative approach, referred to as WindSTORM, to achieve high-speed high-density emitter localization with robust performance for various image characteristics. We demonstrate that WindSTORM improves the computation speed by two orders of magnitude on CPU and three orders of magnitude upon GPU acceleration to realize online image processing, without compromising localization accuracy. Further, due to the embedded background correction, WindSTORM is highly robust in the presence of high and non-uniform background. WindSTORM paves the way for next generation of high-throughput nanoscopy.

scholarly journals Divide and Conquer: Real-time maximum likelihood fitting of multiple emitters for super-resolution localization microscopy

2019 ◽  
Author(s):  
Luchang Li ◽  
Bo Xin ◽  
Weibing Kuang ◽  
Zhiwei Zhou ◽  
Zhen-Li Huang
Keyword(s):  
Image Processing ◽  
Real Time ◽  
Super Resolution ◽  
Large Cell ◽  
Field Of View ◽  
Divide And Conquer ◽  
Image Size ◽  
Localization Microscopy ◽  
Localization Algorithms ◽  
Emitter Localization

AbstractMulti-emitter localization has great potential for maximizing the imaging speed of super-resolution localization microscopy. However, the slow image analysis speed of reported multi-emitter localization algorithms limits their usage in mostly off-line image processing with small image size. Here we adopt the well-known divide and conquer strategy in computer science and present a fitting-based method called QC-STORM for fast multi-emitter localization. Using simulated and experimental data, we verify that QC-STORM is capable of providing real-time full image processing on raw images with 100 µm × 100 µm field of view and 10 ms exposure time, with comparable spatial resolution as the popular fitting-based ThunderSTORM and the up-to-date non-iterative WindSTORM. This study pushes the development and practical use of super-resolution localization microscopy in high-throughput or high-content imaging of cell-to-cell differences or discovering rare events in a large cell population.

scholarly journals Comparing Super-Resolution Microscopy Techniques to Analyze Chromosomes

2021 ◽  
Vol 22 (4) ◽  
pp. 1903
Author(s):  
Ivona Kubalová ◽  
Alžběta Němečková ◽  
Klaus Weisshart ◽  
Eva Hřibová ◽  
Veit Schubert
Keyword(s):  
Single Molecule ◽  
Imaging Techniques ◽  
Chromatin Condensation ◽  
Super Resolution ◽  
Stimulated Emission ◽  
Wide Field ◽  
Structured Illumination Microscopy ◽  
Metaphase Chromosomes ◽  
Localization Microscopy ◽  
Super Resolution Microscopy

The importance of fluorescence light microscopy for understanding cellular and sub-cellular structures and functions is undeniable. However, the resolution is limited by light diffraction (~200–250 nm laterally, ~500–700 nm axially). Meanwhile, super-resolution microscopy, such as structured illumination microscopy (SIM), is being applied more and more to overcome this restriction. Instead, super-resolution by stimulated emission depletion (STED) microscopy achieving a resolution of ~50 nm laterally and ~130 nm axially has not yet frequently been applied in plant cell research due to the required specific sample preparation and stable dye staining. Single-molecule localization microscopy (SMLM) including photoactivated localization microscopy (PALM) has not yet been widely used, although this nanoscopic technique allows even the detection of single molecules. In this study, we compared protein imaging within metaphase chromosomes of barley via conventional wide-field and confocal microscopy, and the sub-diffraction methods SIM, STED, and SMLM. The chromosomes were labeled by DAPI (4′,6-diamidino-2-phenylindol), a DNA-specific dye, and with antibodies against topoisomerase IIα (Topo II), a protein important for correct chromatin condensation. Compared to the diffraction-limited methods, the combination of the three different super-resolution imaging techniques delivered tremendous additional insights into the plant chromosome architecture through the achieved increased resolution.

scholarly journals Camera-based localization microscopy optimized with calibrated structured illumination

2021 ◽  
Vol 4 (1) ◽  
Author(s):  
Martin Schmidt ◽  
Adam C. Hundahl ◽  
Henrik Flyvbjerg ◽  
Rodolphe Marie ◽  
Kim I. Mortensen
Keyword(s):  
Data Analysis ◽  
Super Resolution ◽  
Wide Field ◽  
Structured Illumination ◽  
Uniform Illumination ◽  
Localization Microscopy ◽  
Localization And Tracking ◽  
Field Imaging ◽  
Wide Field Imaging ◽  
Localization Information

AbstractUntil very recently, super-resolution localization and tracking of fluorescent particles used camera-based wide-field imaging with uniform illumination. Then it was demonstrated that structured illuminations encode additional localization information in images. The first demonstration of this uses scanning and hence suffers from limited throughput. This limitation was mitigated by fusing camera-based localization with wide-field structured illumination. Current implementations, however, use effectively only half the localization information that they encode in images. Here we demonstrate how all of this information may be exploited by careful calibration of the structured illumination. Our approach achieves maximal resolution for given structured illumination, has a simple data analysis, and applies to any structured illumination in principle. We demonstrate this with an only slightly modified wide-field microscope. Our protocol should boost the emerging field of high-precision localization with structured illumination.

scholarly journals Optimizing Imaging Conditions for Demanding Multi-Color Super Resolution Localization Microscopy

PLoS ONE ◽  
2016 ◽  
Vol 11 (7) ◽  
pp. e0158884 ◽  
Author(s):  
Leila Nahidiazar ◽  
Alexandra V. Agronskaia ◽  
Jorrit Broertjes ◽  
Bram van den Broek ◽  
Kees Jalink
Keyword(s):  
Super Resolution ◽  
Localization Microscopy ◽  
Imaging Conditions
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