Revealing dynamically-organized receptor ion channel clusters in live cells by a correlated electric recording and super-resolution single-molecule imaging approach

2018 ◽  
Vol 20 (12) ◽  
pp. 8088-8098 ◽  
Author(s):  
Rajeev Yadav ◽  
H. Peter Lu
Keyword(s):  
Ion Channel ◽  
Single Molecule ◽  
Super Resolution ◽  
Live Cells ◽  
Imaging Approach ◽  
Resolution Imaging ◽  
Function Relationship ◽  
Bleaching Step ◽  
Relationship Of ◽  
Resolution Single

Correlating single-molecule fluorescence photo-bleaching step analysis and single-molecule super-resolution imaging, our findings for the clustering effect of the NMDA receptor ion channel on the live cell membranes provide a new and significant understanding of the structure–function relationship of NMDA receptors.

scholarly journals Single cell, super-resolution imaging reveals an acid pH-dependent conformational switch in SsrB regulates SPI-2

eLife ◽  
2019 ◽  
Vol 8 ◽  
Author(s):  
Andrew Tze Fui Liew ◽  
Yong Hwee Foo ◽  
Yunfeng Gao ◽  
Parisa Zangoui ◽  
Moirangthem Kiran Singh ◽  
...  
Keyword(s):  
Dna Binding ◽  
Single Molecule ◽  
Single Cells ◽  
Super Resolution ◽  
Single Particle Tracking ◽  
Live Cells ◽  
Bacterial Cells ◽  
Acid Ph ◽  
Resolution Imaging ◽  
Ph Dependent

After Salmonella is phagocytosed, it resides in an acidic vacuole. Its cytoplasm acidifies to pH 5.6; acidification activates pathogenicity island 2 (SPI-2). SPI-2 encodes a type three secretion system whose effectors modify the vacuole, driving endosomal tubulation. Using super-resolution imaging in single bacterial cells, we show that low pH induces expression of the SPI-2 SsrA/B signaling system. Single particle tracking, atomic force microscopy, and single molecule unzipping assays identified pH-dependent stimulation of DNA binding by SsrB. A so-called phosphomimetic form (D56E) was unable to bind to DNA in live cells. Acid-dependent DNA binding was not intrinsic to regulators, as PhoP and OmpR binding was not pH-sensitive. The low level of SPI-2 injectisomes observed in single cells is not due to fluctuating SsrB levels. This work highlights the surprising role that acid pH plays in virulence and intracellular lifestyles of Salmonella; modifying acid survival pathways represents a target for inhibiting Salmonella.

scholarly journals A H-bond strategy to develop acid-resistant photoswitchable rhodamine spirolactams for super-resolution single-molecule localization microscopy

2019 ◽  
Vol 10 (18) ◽  
pp. 4914-4922 ◽  
Author(s):  
Qingkai Qi ◽  
Weijie Chi ◽  
Yuanyuan Li ◽  
Qinglong Qiao ◽  
Jie Chen ◽  
...  
Keyword(s):  
Single Molecule ◽  
Super Resolution ◽  
Amino Groups ◽  
Localization Microscopy ◽  
Resolution Imaging ◽  
Resolution Single ◽  
Single Molecule Localization Microscopy

Rhodamine spirolactams with adjacent amino groups work as acid-resistant and photoswitchable fluorophores in single-molecule localization super-resolution imaging.

scholarly journals High Numerical Aperture Epi-illumination Selective Plane Illumination Microscopy

2018 ◽  
Author(s):  
Bin Yang ◽  
Yina Wang ◽  
Siyu Feng ◽  
Veronica Pessino ◽  
Nico Stuurman ◽  
...  
Keyword(s):  
Single Molecule ◽  
Numerical Aperture ◽  
Super Resolution ◽  
Live Cells ◽  
Volumetric Imaging ◽  
High Numerical Aperture ◽  
Selective Plane Illumination Microscopy ◽  
Resolution Imaging ◽  
Living Organisms ◽  
Super Resolution Microscopy

Selective-plane illumination microscopy (SPIM) provides unparalleled advantages for long-term volumetric imaging of living organisms. In order to achieve high-resolution imaging in common biological sample holders, we designed a high numerical aperture (NA) epi-illumination SPIM (eSPIM) system, which utilizes a single objective and has an identical sample interface as an inverted fluorescence microscope with no additional reflection elements. This system has an effective detection NA of > 1.06. We demonstrated multicolor and fast volumetric imaging of live cells and single-molecule super-resolution microscopy using our system.

scholarly journals 3D super-resolved imaging in live cells using sub-diffractive plasmonic localization of hybrid nanopillar arrays

Nanophotonics ◽  
2020 ◽  
Vol 9 (9) ◽  
pp. 2847-2859
Author(s):  
Soojung Kim ◽  
Hyerin Song ◽  
Heesang Ahn ◽  
Seung Won Jun ◽  
Seungchul Kim ◽  
...  
Keyword(s):  
Signal To Noise Ratio ◽  
Imaging Techniques ◽  
Optical Loss ◽  
Super Resolution ◽  
Living Cells ◽  
Live Cells ◽  
Complex Biological System ◽  
Observation Volume ◽  
Resolution Imaging ◽  
Nanopillar Arrays

AbstractAnalysing dynamics of a single biomolecule using high-resolution imaging techniques has been had significant attentions to understand complex biological system. Among the many approaches, vertical nanopillar arrays in contact with the inside of cells have been reported as a one of useful imaging applications since an observation volume can be confined down to few-tens nanometre theoretically. However, the nanopillars experimentally are not able to obtain super-resolution imaging because their evanescent waves generate a high optical loss and a low signal-to-noise ratio. Also, conventional nanopillars have a limitation to yield 3D information because they do not concern field localization in z-axis. Here, we developed novel hybrid nanopillar arrays (HNPs) that consist of SiO2 nanopillars terminated with gold nanodisks, allowing extreme light localization. The electromagnetic field profiles of HNPs are obtained through simulations and imaging resolution of cell membrane and biomolecules in living cells are tested using one-photon and 3D multiphoton fluorescence microscopy, respectively. Consequently, HNPs present approximately 25 times enhanced intensity compared to controls and obtained an axial and lateral resolution of 110 and 210 nm of the intensities of fluorophores conjugated with biomolecules transported in living cells. These structures can be a great platform to analyse complex intracellular environment.

scholarly journals Comparing lifeact and phalloidin for super-resolution imaging of actin in fixed cells

PLoS ONE ◽  
2021 ◽  
Vol 16 (1) ◽  
pp. e0246138
Author(s):  
Hanieh Mazloom-Farsibaf ◽  
Farzin Farzam ◽  
Mohamadreza Fazel ◽  
Michael J. Wester ◽  
Marjolein B. M. Meddens ◽  
...  
Keyword(s):  
Single Molecule ◽  
Cell Biology ◽  
Cell Movement ◽  
Super Resolution ◽  
Actin Binding ◽  
Thin Filaments ◽  
Reversible Binding ◽  
Multiple Regions ◽  
Resolution Imaging ◽  
Division Cell

Visualizing actin filaments in fixed cells is of great interest for a variety of topics in cell biology such as cell division, cell movement, and cell signaling. We investigated the possibility of replacing phalloidin, the standard reagent for super-resolution imaging of F-actin in fixed cells, with the actin binding peptide ‘lifeact’. We compared the labels for use in single molecule based super-resolution microscopy, where AlexaFluor 647 labeled phalloidin was used in a dSTORM modality and Atto 655 labeled lifeact was used in a single molecule imaging, reversible binding modality. We found that imaging with lifeact had a comparable resolution in reconstructed images and provided several advantages over phalloidin including lower costs, the ability to image multiple regions of interest on a coverslip without degradation, simplified sequential super-resolution imaging, and more continuous labeling of thin filaments.

Single molecule Coordinate and Height super-resolution Imaging with Dithering and Orientation (CHIDO)

2020 ◽  
Author(s):  
Luis A. Alemán-Castañeda ◽  
Valentina Curcio ◽  
Thomas G. Brown ◽  
Sophie Brasselet ◽  
Miguel A. Alonso
Keyword(s):  
Single Molecule ◽  
Super Resolution ◽  
Resolution Imaging

scholarly journals Fast, Simultaneous Multiple Fluorophore Fitting in Single Molecule Super-Resolution Imaging

2011 ◽  
Vol 100 (3) ◽  
pp. 349a
Author(s):  
Fang Huang ◽  
Samantha L. Schwartz ◽  
Jason M. Byars ◽  
Keith A. Lidke
Keyword(s):  
Single Molecule ◽  
Super Resolution ◽  
Resolution Imaging ◽  
Fitting In
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