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Cells
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Development and Challenges in Microscopy for Cellular Imaging
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Development and Challenges in Microscopy for Cellular Imaging

A special issue of Cells (ISSN 2073-4409). This special issue belongs to the section "Cell Nuclei: Function, Transport and Receptors".

Deadline for manuscript submissions: closed (31 May 2019) | Viewed by 44058

Special Issue Editor

Prof. Dr. Ohad Medalia

E-Mail Website
Guest Editor
1. Department of Biochemistry, University of Zurich, Zurich, Switzerland
2. Department of Life Sciences and the National Institute for Biotechnology in the Negev, Ben-Gurion University, Beer-Sheva, Israel
Interests: nuclear envelope; lamins; cell adhesion; cryo-electron tomography
Special Issues, Collections and Topics in MDPI journals

Special Issue Information

Dear Colleagues,

Cellular imaging and structural approaches have long since revolutionized our knowledge on cellular processes. However, over the last decade we have witnessed tremendous technical advances that challenged earlier limitations and views on cellular imaging. Fluorescence imaging has evolved into an analytical and quantitative tool to reach an accuracy of several manometers in localizing fluorophores. Cryo-EM and tomography allow viewing the molecular architecture of cells at close to life state, opening a window of opportunity towards cellular structural biology. Nuclear magnetic resonance, single molecule and force spectroscopy complement cellular imaging to produce a realistic model of cells in health and disease. Further, multicellular organs and even organisms may now be imaged at the cellular level, revealing a three-dimensional architecture that was previously difficult to obtain.

In this Special Issue of Cells, we invite contributions, in the form of either original research articles or reviews, on aspects related to Cellular Imaging; Papers concerning both technical issues or applications are welcome.

Prof. Ohad Medalia
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. Cells is an international peer-reviewed open access semimonthly journal published by MDPI.

Please visit the Instructions for Authors page before submitting a manuscript. The Article Processing Charge (APC) for publication in this open access journal is 2700 CHF (Swiss Francs). Submitted papers should be well formatted and use good English. Authors may use MDPI's English editing service prior to publication or during author revisions.

Keywords

  • Atomic force microscopy
  • Electron microscopy
  • Fluorescent microscopy
  • Live cell imaging
  • Nuclear magnetic resonance of cells and organelles

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Published Papers (5 papers)

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Research

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12 pages, 6621 KiB  
Article
Noninvasive Subcellular Imaging Using Atomic Force Acoustic Microscopy (AFAM)
by Xiaoqing Li, Ang Lu, Wenjie Deng, Li Su, Jing Wang and Mingyue Ding
Cells 2019, 8(4), 314; https://doi.org/10.3390/cells8040314 - 5 Apr 2019
Cited by 6 | Viewed by 4409
Abstract
We report an imaging approach applying the atomic force acoustic microscopy (AFAM), which has unique potential for nondestructive imaging of cell internal structures. To obtain high spatial resolution images, we optimized the significant imaging parameters, including scanning speeds, feedback configurations and acoustic frequencies [...] Read more.
We report an imaging approach applying the atomic force acoustic microscopy (AFAM), which has unique potential for nondestructive imaging of cell internal structures. To obtain high spatial resolution images, we optimized the significant imaging parameters, including scanning speeds, feedback configurations and acoustic frequencies of an AFAM system, to increase the amplitude of the acoustic signal and to stabilize the morphological signals. We also combined the acoustic amplitude and phase signals, and generated pseudo-color figures for better illustration of subcellular features such as pseudopodia, membranes and nucleus-like. The subcellular structural image atlas can describe nanoscale details of multiple samples and provide clearer images of the subcellular features compared to other conventional techniques. This study builds a strong basis of transmission AFAM for cell imaging, which can help researchers to clarify the cell structures in diverse biological fields and push the understanding of biology evolution to a new stage. Full article
(This article belongs to the Special Issue Development and Challenges in Microscopy for Cellular Imaging)
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20 pages, 5241 KiB  
Article
Quantitative Distribution of DNA, RNA, Histone and Proteins Other than Histone in Mammalian Cells, Nuclei and a Chromosome at High Resolution Observed by Scanning Transmission Soft X-Ray Microscopy (STXM)
by Kunio Shinohara, Shigenobu Toné, Takeo Ejima, Takuji Ohigashi and Atsushi Ito
Cells 2019, 8(2), 164; https://doi.org/10.3390/cells8020164 - 16 Feb 2019
Cited by 9 | Viewed by 4571
Abstract
Soft X-ray microscopy was applied to study the quantitative distribution of DNA, RNA, histone, and proteins other than histone (represented by BSA) in mammalian cells, apoptotic nuclei, and a chromosome at spatial resolutions of 100 to 400 nm. The relative distribution of closely [...] Read more.
Soft X-ray microscopy was applied to study the quantitative distribution of DNA, RNA, histone, and proteins other than histone (represented by BSA) in mammalian cells, apoptotic nuclei, and a chromosome at spatial resolutions of 100 to 400 nm. The relative distribution of closely related molecules, such as DNA and RNA, was discriminated by the singular value decomposition (SVD) method using aXis2000 software. Quantities of nucleic acids and proteins were evaluated using characteristic absorption properties due to the 1s–π * transition of N=C in nucleic acids and amide in proteins, respectively, in the absorption spectra at the nitrogen K absorption edge. The results showed that DNA and histone were located in the nucleus. By contrast, RNA was clearly discriminated and found mainly in the cytoplasm. Interestingly, in a chromosome image, DNA and histone were found in the center, surrounded by RNA and proteins other than histone. The amount of DNA in the chromosome was estimated to be 0.73 pg, and the content of RNA, histone, and proteins other than histone, relative to DNA, was 0.48, 0.28, and 4.04, respectively. The method we present in this study could be a powerful approach for the quantitative molecular mapping of biological samples at high resolution. Full article
(This article belongs to the Special Issue Development and Challenges in Microscopy for Cellular Imaging)
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13 pages, 2126 KiB  
Article
Nanobody Detection of Standard Fluorescent Proteins Enables Multi-Target DNA-PAINT with High Resolution and Minimal Displacement Errors
by Shama Sograte-Idrissi, Nazar Oleksiievets, Sebastian Isbaner, Mariana Eggert-Martinez, Jörg Enderlein, Roman Tsukanov and Felipe Opazo
Cells 2019, 8(1), 48; https://doi.org/10.3390/cells8010048 - 14 Jan 2019
Cited by 43 | Viewed by 9474
Abstract
DNA point accumulation for imaging in nanoscale topography (PAINT) is a rapidly developing fluorescence super-resolution technique, which allows for reaching spatial resolutions below 10 nm. It also enables the imaging of multiple targets in the same sample. However, using DNA-PAINT to observe cellular [...] Read more.
DNA point accumulation for imaging in nanoscale topography (PAINT) is a rapidly developing fluorescence super-resolution technique, which allows for reaching spatial resolutions below 10 nm. It also enables the imaging of multiple targets in the same sample. However, using DNA-PAINT to observe cellular structures at such resolution remains challenging. Antibodies, which are commonly used for this purpose, lead to a displacement between the target protein and the reporting fluorophore of 20–25 nm, thus limiting the resolving power. Here, we used nanobodies to minimize this linkage error to ~4 nm. We demonstrate multiplexed imaging by using three nanobodies, each able to bind to a different family of fluorescent proteins. We couple the nanobodies with single DNA strands via a straight forward and stoichiometric chemical conjugation. Additionally, we built a versatile computer-controlled microfluidic setup to enable multiplexed DNA-PAINT in an efficient manner. As a proof of principle, we labeled and imaged proteins on mitochondria, the Golgi apparatus, and chromatin. We obtained super-resolved images of the three targets with 20 nm resolution, and within only 35 minutes acquisition time. Full article
(This article belongs to the Special Issue Development and Challenges in Microscopy for Cellular Imaging)
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Review

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20 pages, 6829 KiB  
Review
Quantitative Analysis of Nuclear Lamins Imaged by Super-Resolution Light Microscopy
by Mark Kittisopikul, Laura Virtanen, Pekka Taimen and Robert D. Goldman
Cells 2019, 8(4), 361; https://doi.org/10.3390/cells8040361 - 18 Apr 2019
Cited by 13 | Viewed by 6178
Abstract
The nuclear lamina consists of a dense fibrous meshwork of nuclear lamins, Type V intermediate filaments, and is ~14 nm thick according to recent cryo-electron tomography studies. Recent advances in light microscopy have extended the resolution to a scale allowing for the fine [...] Read more.
The nuclear lamina consists of a dense fibrous meshwork of nuclear lamins, Type V intermediate filaments, and is ~14 nm thick according to recent cryo-electron tomography studies. Recent advances in light microscopy have extended the resolution to a scale allowing for the fine structure of the lamina to be imaged in the context of the whole nucleus. We review quantitative approaches to analyze the imaging data of the nuclear lamina as acquired by structured illumination microscopy (SIM) and single molecule localization microscopy (SMLM), as well as the requisite cell preparation techniques. In particular, we discuss the application of steerable filters and graph-based methods to segment the structure of the four mammalian lamin isoforms (A, C, B1, and B2) and extract quantitative information. Full article
(This article belongs to the Special Issue Development and Challenges in Microscopy for Cellular Imaging)
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18 pages, 10034 KiB  
Review
Cellular and Structural Studies of Eukaryotic Cells by Cryo-Electron Tomography
by Miriam Sarah Weber, Matthias Wojtynek and Ohad Medalia
Cells 2019, 8(1), 57; https://doi.org/10.3390/cells8010057 - 16 Jan 2019
Cited by 39 | Viewed by 18528
Abstract
The architecture of protein assemblies and their remodeling during physiological processes is fundamental to cells. Therefore, providing high-resolution snapshots of macromolecular complexes in their native environment is of major importance for understanding the molecular biology of the cell. Cellular structural biology by means [...] Read more.
The architecture of protein assemblies and their remodeling during physiological processes is fundamental to cells. Therefore, providing high-resolution snapshots of macromolecular complexes in their native environment is of major importance for understanding the molecular biology of the cell. Cellular structural biology by means of cryo-electron tomography (cryo-ET) offers unique insights into cellular processes at an unprecedented resolution. Recent technological advances have enabled the detection of single impinging electrons and improved the contrast of electron microscopic imaging, thereby significantly increasing the sensitivity and resolution. Moreover, various sample preparation approaches have paved the way to observe every part of a eukaryotic cell, and even multicellular specimens, under the electron beam. Imaging of macromolecular machineries at high resolution directly within their native environment is thereby becoming reality. In this review, we discuss several sample preparation and labeling techniques that allow the visualization and identification of macromolecular assemblies in situ, and demonstrate how these methods have been used to study eukaryotic cellular landscapes. Full article
(This article belongs to the Special Issue Development and Challenges in Microscopy for Cellular Imaging)
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