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Development and Application of Super-resolution Imaging Methods for Biological Research 2.0

A special issue of International Journal of Molecular Sciences (ISSN 1422-0067). This special issue belongs to the section "Molecular Biophysics".

Deadline for manuscript submissions: closed (30 November 2022) | Viewed by 9903

Special Issue Editor

Dr. Kiryl Piatkevich

E-Mail Website
Guest Editor
1. McGovern Institute for Brain Research, Cambridge, MA, USA
2. Molecular BioEngineering Group, School of Life Sciences, Westlake University, Hangzhou, China
Interests: fluorescent biosensors; optogenetics; expension microscop; neuroengineering
Special Issues, Collections and Topics in MDPI journals

Special Issue Information

Dear Colleagues,

Biological research and biomedicine are increasingly reliant on optical methods for imaging the structure and function of biological systems. Addressing the most pressing needs in biological research requires visualization and mapping biomolecules with nanoscale precision. Recent advances in optical techniques in combination with the development of novel molecular probes made it possible to break the diffraction limit of conventional optical microscopy, establishing super-resolution microscopy, for which the 2014 Nobel Prize in Chemistry was awarded. The field of super-resolution microscopy remains extremely dynamic and fast-growing and new methods and techniques are emerging all the time.

We would like to kindly invite you to contribute to the Special Issue "Development and Application of Super-Resolution Imaging Methods for Biological Research" of the International Journal of Molecular Sciences (Impact Factor 4.556). The Special Issue will elucidate the development and application of new imaging methods and techniques that enable sub-diffraction imaging of biological systems. Novel molecular probes and sample preparation techniques that enhance existing super-resolution methods or provide new modalities in super-resolution imaging are also within the scope of the Special Issue. Papers may also focus on addressing important biological questions using super-resolution imaging techniques. The Special Issue will provide a forum for sharing and discussing perspectives on the development of the field within biological research. Review articles can cover recent advances in the field of super-resolution microscopy or molecular probes for super-resolution microscopy, or novel or emerging techniques for sub-diffraction imaging.

Dr. Kiryl Piatkevich
Guest Editor

Manuscript Submission Information

Manuscripts should be submitted online at www.mdpi.com by registering and logging in to this website. Once you are registered, click here to go to the submission form. Manuscripts can be submitted until the deadline. All submissions that pass pre-check are peer-reviewed. Accepted papers will be published continuously in the journal (as soon as accepted) and will be listed together on the special issue website. Research articles, review articles as well as short communications are invited. For planned papers, a title and short abstract (about 100 words) can be sent to the Editorial Office for announcement on this website.

Submitted manuscripts should not have been published previously, nor be under consideration for publication elsewhere (except conference proceedings papers). All manuscripts are thoroughly refereed through a single-blind peer-review process. A guide for authors and other relevant information for submission of manuscripts is available on the Instructions for Authors page. International Journal of Molecular Sciences is an international peer-reviewed open access semimonthly journal published by MDPI.

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Keywords

  • 4Pi microscopy
  • STED and two-photon STED
  • RESOLFT
  • localization microscopy
  • structured illumination microscopy
  • super-resolution optical fluctuation imaging
  • light-sheet microscopy
  • expansion microscopy
  • light-induced photoconversion
  • application of photostable emitters
  • tissue clearing
  • sample magnification via isotropic expansion
  • multiplexed staining
  • fluorescent proteins
  • advanced antibodies and nanobodies
  • novel or special dyes
  • new instrumentation and new techniques
  • enhanced resolution
  • in vivo application
  • large sample imaging

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Published Papers (3 papers)

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Research

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13 pages, 10873 KiB  
Article
Long Preservation of AAV-Transduced Fluorescence by a Modified Organic Solvent-Based Clearing Method
by Tao Lu, Munehisa Shinozaki, Narihito Nagoshi, Masaya Nakamura and Hideyuki Okano
Int. J. Mol. Sci. 2022, 23(17), 9637; https://doi.org/10.3390/ijms23179637 - 25 Aug 2022
Cited by 1 | Viewed by 2182
Abstract
The development of tissue clearing technologies allows 3D imaging of whole tissues and organs, especially in studies of the central nervous system innervated throughout the body. Although the three-dimensional imaging of solvent-cleared organs (3DISCO) method provides a powerful clearing capacity and high transparency, [...] Read more.
The development of tissue clearing technologies allows 3D imaging of whole tissues and organs, especially in studies of the central nervous system innervated throughout the body. Although the three-dimensional imaging of solvent-cleared organs (3DISCO) method provides a powerful clearing capacity and high transparency, the rapid quenching of endogenous fluorescence and peroxide removal process decreases its practicability. This study provides a modified method named tDISCO to solve these limitations. The tDISCO protocol can preserve AAV-transduced endogenous EGFP fluorescence for months and achieve high transparency in a fast and simple clearing process. In addition to the brain, tDISCO was applied to other organs and even hard bone tissue. tDISCO also enabled us to visualize the long projection neurons and axons with high resolution. This method provides a fast and simple clearing protocol for 3D visualization of the AAV- transduced long projection neurons throughout the brain and spinal cord. Full article
(This article belongs to the Special Issue Development and Application of Super-resolution Imaging Methods for Biological Research 2.0)
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18 pages, 4220 KiB  
Article
CRISPR/Cas9 Genome Editing vs. Over-Expression for Fluorescent Extracellular Vesicle-Labeling: A Quantitative Analysis
by Karin Strohmeier, Martina Hofmann, Fabian Hauser, Dmitry Sivun, Sujitha Puthukodan, Andreas Karner, Georg Sandner, Pol-Edern Le Renard, Jaroslaw Jacak and Mario Mairhofer
Int. J. Mol. Sci. 2022, 23(1), 282; https://doi.org/10.3390/ijms23010282 - 28 Dec 2021
Cited by 10 | Viewed by 3858
Abstract
Over-expression of fluorescently-labeled markers for extracellular vesicles is frequently used to visualize vesicle up-take and transport. EVs that are labeled by over-expression show considerable heterogeneity regarding the number of fluorophores on single particles, which could potentially bias tracking and up-take studies in favor [...] Read more.
Over-expression of fluorescently-labeled markers for extracellular vesicles is frequently used to visualize vesicle up-take and transport. EVs that are labeled by over-expression show considerable heterogeneity regarding the number of fluorophores on single particles, which could potentially bias tracking and up-take studies in favor of more strongly-labeled particles. To avoid the potential artefacts that are caused by over-expression, we developed a genome editing approach for the fluorescent labeling of the extracellular vesicle marker CD63 with green fluorescent protein using the CRISPR/Cas9 technology. Using single-molecule sensitive fluorescence microscopy, we quantitatively compared the degree of labeling of secreted small extracellular vesicles from conventional over-expression and the CRISPR/Cas9 approach with true single-particle measurements. With our analysis, we can demonstrate a larger fraction of single-GFP-labeled EVs in the EVs that were isolated from CRISPR/Cas9-modified cells (83%) compared to EVs that were isolated from GFP-CD63 over-expressing cells (36%). Despite only single-GFP-labeling, CRISPR-EVs can be detected and discriminated from auto-fluorescence after their up-take into cells. To demonstrate the flexibility of the CRISPR/Cas9 genome editing method, we fluorescently labeled EVs using the HaloTag® with lipid membrane permeable dye, JaneliaFluor® 646, which allowed us to perform 3D-localization microscopy of single EVs taken up by the cultured cells. Full article
(This article belongs to the Special Issue Development and Application of Super-resolution Imaging Methods for Biological Research 2.0)
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Review

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18 pages, 3077 KiB  
Review
Transient Fluorescence Labeling: Low Affinity—High Benefits
by Maxim M. Perfilov, Alexey S. Gavrikov, Konstantin A. Lukyanov and Alexander S. Mishin
Int. J. Mol. Sci. 2021, 22(21), 11799; https://doi.org/10.3390/ijms222111799 - 30 Oct 2021
Cited by 7 | Viewed by 3197
Abstract
Fluorescent labeling is an established method for visualizing cellular structures and dynamics. The fundamental diffraction limit in image resolution was recently bypassed with the development of super-resolution microscopy. Notably, both localization microscopy and stimulated emission depletion (STED) microscopy impose tight restrictions on the [...] Read more.
Fluorescent labeling is an established method for visualizing cellular structures and dynamics. The fundamental diffraction limit in image resolution was recently bypassed with the development of super-resolution microscopy. Notably, both localization microscopy and stimulated emission depletion (STED) microscopy impose tight restrictions on the physico-chemical properties of labels. One of them—the requirement for high photostability—can be satisfied by transiently interacting labels: a constant supply of transient labels from a medium replenishes the loss in the signal caused by photobleaching. Moreover, exchangeable tags are less likely to hinder the intrinsic dynamics and cellular functions of labeled molecules. Low-affinity labels may be used both for fixed and living cells in a range of nanoscopy modalities. Nevertheless, the design of optimal labeling and imaging protocols with these novel tags remains tricky. In this review, we highlight the pros and cons of a wide variety of transiently interacting labels. We further discuss the state of the art and future perspectives of low-affinity labeling methods. Full article
(This article belongs to the Special Issue Development and Application of Super-resolution Imaging Methods for Biological Research 2.0)
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