Super-resolution microscopy (PALM, STORM etc.) provides a plethora of fluorescent signals in dense cellular environments which can be difficult to interpret. Here we describe ClusterViSu, a method for image reconstruction, visualization and quantification of labelled protein clusters, based on Voronoi tessellation of the individual fluorescence events. The general applicability of this clustering approach for the segmentation of super-resolution microscopy data, including for co-localization, is illustrated on a series of important biological objects such as chromatin complexes, RNA polymerase, nuclear pore complexes and microtubules.
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http://dx.doi.org/10.1038/srep24084 | DOI Listing |
Optica
June 2021
Department of Electrical Engineering, University of Notre Dame, Notre Dame, IN 46556, USA.
Traditional fluorescence microscopy is blind to molecular microenvironment information that is present in fluorescence lifetime, which can be measured by fluorescence lifetime imaging microscopy (FLIM). However, most existing FLIM techniques are slow to acquire and process lifetime images, difficult to implement, and expensive. Here, we present instant FLIM, an analog signal processing method that allows real-time streaming of fluorescence intensity, lifetime, and phasor imaging data through simultaneous image acquisition and instantaneous data processing.
View Article and Find Full Text PDFJ Cell Sci
January 2025
Department of Cellular & Physiological Sciences, Life Sciences Institute, University of British Columbia, Vancouver, BC V6T 1Z3, Canada.
Here, we apply SuperResNET network analysis of dSTORM single-molecule localization microscopy (SMLM) to determine how the clathrin endocytosis inhibitors pitstop 2, dynasore and Latrunculin A alter the morphology of clathrin-coated pits. SuperResNET analysis of HeLa and Cos7 cells identifies: small oligomers (Class I); pits and vesicles (Class II); and larger clusters corresponding to fused pits or clathrin plaques (Class III). Pitstop 2 and dynasore induce distinct homogeneous populations of Class II structures in HeLa cells suggesting that they arrest endocytosis at different stages.
View Article and Find Full Text PDFAnal Chem
January 2025
Shandong Provincial Key Laboratory of Tumor Imaging Equipment Development and Integrated Diagnosis and Treatment Technology, Linyi University, Linyi 276000, China.
Drug-induced liver injury (DILI) has emerged as an urgent clinical challenge. It is characterized by mitochondrial dysfunction in liver cells, which leads to abnormal changes in HO levels within the mitochondria. Super-resolution imaging allows for the observation of the fine structure of mitochondria at the nanometer scale, potentially enabling the detection of mitochondrial HO levels during DILI at the subcellular organelle level.
View Article and Find Full Text PDFProtein Sci
February 2025
IRR Chemistry Hub, Institute for Regeneration and Repair, The University of Edinburgh, Edinburgh, UK.
Super-resolution microscopy has revolutionized biological imaging, enabling the visualization of structures at the nanometer length scale. Its application in live cells, however, has remained challenging. To address this, we adapted LIVE-PAINT, an approach we established in yeast, for application in live mammalian cells.
View Article and Find Full Text PDFCell Chem Biol
January 2025
Yusuf Hamied Department of Chemistry, University of Cambridge Cambridge CB2 1EW, UK; UK Dementia Research Institute at University of Cambridge Cambridge CB2 0XY, UK. Electronic address:
Synaptic dysfunction is a primary hallmark of both Alzheimer's and Parkinson's disease, leading to cognitive and behavioral decline. While alpha-synuclein, beta-amyloid, and tau are involved in the physiological functioning of synapses, their pathological aggregation has been linked to synaptopathology. The methodology for studying the small-soluble protein aggregates formed by these proteins is limited.
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