scholarly journals Ultra-fast super-resolution imaging of biomolecular mobility in tissues

2017 ◽  
Author(s):  
Helen Miller ◽  
Jason Cosgrove ◽  
Adam J. M. Wollman ◽  
Peter O’ J. Toole ◽  
Mark C. Coles ◽  
...  
Keyword(s):  
Lymph Node ◽  
Light Microscopy ◽  
Molecular Diffusion ◽  
Ex Vivo ◽  
Super Resolution ◽  
Collagen Matrix ◽  
Tissue Sections ◽  
Aqueous Environments ◽  
Resolution Imaging

Super-resolution techniques have addressed many biological questions, yet molecular quantification at rapid timescales in live tissues remains challenging. We developed a light microscopy system capable of sub-millisecond sampling to characterize molecular diffusion in heterogeneous aqueous environments comparable to interstitial regions between cells in tissues. We demonstrate our technique with super-resolution tracking of fluorescently labelled chemokine molecules in a collagen matrix andex vivolymph node tissue sections, outperforming competing methods.

Plasmonics-based spatially activated light microscopy for super-resolution imaging of molecular fluorescence

2010 ◽  
Vol 35 (20) ◽  
pp. 3501 ◽  
Author(s):  
Kyujung Kim ◽  
Youngjin Oh ◽  
Wonju Lee ◽  
Donghyun Kim
Keyword(s):  
Light Microscopy ◽  
Super Resolution ◽  
Molecular Fluorescence ◽  
Resolution Imaging

scholarly journals How To Extend The Capabilities Of A Commercial Two-Photon Microscope To Perform Super-Resolution Imaging, Wavelength Mixing And Label-Free Microscopy.

2021 ◽  
Author(s):  
Chiara Peres ◽  
Chiara Nardin ◽  
Guang Yang ◽  
Fabio Mammano
Keyword(s):  
Ex Vivo ◽  
Super Resolution ◽  
Label Free ◽  
Linear Optics ◽  
Non Linear Optics ◽  
Two Photon ◽  
Custom Made ◽  
Non Linear ◽  
Resolution Imaging

Multimodal microscopy combines multiple non-linear techniques that take advantage of different optical processes to generate contrast and increase the amount of information that can be obtained from biological samples. However, the most advanced optical architectures are typically custom-made and require complex alignment procedures, as well as daily maintenance by properly trained personnel for optimal performance. Here, we describe a hybrid system we constructed to overcome these disadvantages by modifying a commercial upright microscope. We show that our multimodal imaging platform can be used to seamlessly perform two-photon STED, wavelength mixing and label-free microscopy in both ex vivo and in vivo samples. The system is highly stable and endowed with remote alignment hardware that ensures simplified operability for non-expert users. This optical architecture is an important step forward towards a wider practical applicability of non-linear optics to bioimaging.

scholarly journals Focusing super resolution on the cytoskeleton

F1000Research ◽  
2016 ◽  
Vol 5 ◽  
pp. 998 ◽  
Author(s):  
Eric A. Shelden ◽  
Zachary T. Colburn ◽  
Jonathan C.R. Jones
Keyword(s):  
Light Microscopy ◽  
Super Resolution ◽  
Living Cells ◽  
Specimen Preparation ◽  
Single Specimen ◽  
Dynamic Changes ◽  
Preparation Methods ◽  
New Methods ◽  
Resolution Imaging ◽  
Over Time

Super resolution imaging is becoming an increasingly important tool in the arsenal of methods available to cell biologists. In recognition of its potential, the Nobel Prize for chemistry was awarded to three investigators involved in the development of super resolution imaging methods in 2014. The availability of commercial instruments for super resolution imaging has further spurred the development of new methods and reagents designed to take advantage of super resolution techniques. Super resolution offers the advantages traditionally associated with light microscopy, including the use of gentle fixation and specimen preparation methods, the ability to visualize multiple elements within a single specimen, and the potential to visualize dynamic changes in living specimens over time. However, imaging of living cells over time is difficult and super resolution imaging is computationally demanding. In this review, we discuss the advantages/disadvantages of different super resolution systems for imaging fixed live specimens, with particular regard to cytoskeleton structures.

scholarly journals Fluorescent Protein-Based Indicators for Functional Super-Resolution Imaging of Biomolecular Activities in Living Cells

2019 ◽  
Vol 20 (22) ◽  
pp. 5784 ◽  
Author(s):  
Kai Lu ◽  
Cong Quang Vu ◽  
Tomoki Matsuda ◽  
Takeharu Nagai
Keyword(s):  
Image Analysis ◽  
Light Microscopy ◽  
Fluorescent Protein ◽  
Fluorescent Proteins ◽  
Super Resolution ◽  
Living Cells ◽  
Bimolecular Fluorescence Complementation ◽  
Analysis Pipeline ◽  
Nanoscale Topography ◽  
Resolution Imaging

Super-resolution light microscopy (SRM) offers a unique opportunity for diffraction-unlimited imaging of biomolecular activities in living cells. To realize such potential, genetically encoded indicators were developed recently from fluorescent proteins (FPs) that exhibit phototransformation behaviors including photoactivation, photoconversion, and photoswitching, etc. Super-resolution observations of biomolecule interactions and biochemical activities have been demonstrated by exploiting the principles of bimolecular fluorescence complementation (BiFC), points accumulation for imaging nanoscale topography (PAINT), and fluorescence fluctuation increase by contact (FLINC), etc. To improve functional nanoscopy with the technology of genetically encoded indicators, it is essential to fully decipher the photo-induced chemistry of FPs and opt for innovative indicator designs that utilize not only fluorescence intensity but also multi-parametric readouts such as phototransformation kinetics. In parallel, technical improvements to both the microscopy optics and image analysis pipeline are promising avenues to increase the sensitivity and versatility of functional SRM.

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