scholarly journals Looking back and looking forward: contributions of electron microscopy to the structural cell biology of gametes and fertilization

Open Biology ◽  
2020 ◽  
Vol 10 (9) ◽  
pp. 200186
Author(s):  
Ravi Teja Ravi ◽  
Miguel Ricardo Leung ◽  
Tzviya Zeev-Ben-Mordehai
Keyword(s):  
Electron Microscopy ◽  
Cell Biology ◽  
Assisted Reproductive Technologies ◽  
Electron Tomography ◽  
Reproductive Technologies ◽  
Practical Significance ◽  
Preparation Methods ◽  
Structural Cell ◽  
Levels Of Detail ◽  
Complex Membrane

Mammalian gametes—the sperm and the egg—represent opposite extremes of cellular organization and scale. Studying the ultrastructure of gametes is crucial to understanding their interactions, and how to manipulate them in order to either encourage or prevent their union. Here, we survey the prominent electron microscopy (EM) techniques, with an emphasis on considerations for applying them to study mammalian gametes. We review how conventional EM has provided significant insight into gamete ultrastructure, but also how the harsh sample preparation methods required preclude understanding at a truly molecular level. We present recent advancements in cryo-electron tomography that provide an opportunity to image cells in a near-native state and at unprecedented levels of detail. New and emerging cellular EM techniques are poised to rekindle exploration of fundamental questions in mammalian reproduction, especially phenomena that involve complex membrane remodelling and protein reorganization. These methods will also allow novel lines of enquiry into problems of practical significance, such as investigating unexplained causes of human infertility and improving assisted reproductive technologies for biodiversity conservation.

Frontiers in Cryo Electron Microscopy of Complex Macromolecular Assemblies

2019 ◽  
Vol 21 (1) ◽  
pp. 395-415 ◽  
Author(s):  
Jana Ognjenović ◽  
Reinhard Grisshammer ◽  
Sriram Subramaniam
Keyword(s):  
Electron Microscopy ◽  
Cell Biology ◽  
Electron Tomography ◽  
Protein Complexes ◽  
Specimen Preparation ◽  
Macromolecular Assemblies ◽  
Cryo Electron Microscopy ◽  
G Protein Coupled ◽  
Improved Methods

In recent years, cryo electron microscopy (cryo-EM) technology has been transformed with the development of better instrumentation, direct electron detectors, improved methods for specimen preparation, and improved software for data analysis. Analyses using single-particle cryo-EM methods have enabled determination of structures of proteins with sizes smaller than 100 kDa and resolutions of ∼2 Å in some cases. The use of electron tomography combined with subvolume averaging is beginning to allow the visualization of macromolecular complexes in their native environment in unprecedented detail. As a result of these advances, solutions to many intractable challenges in structural and cell biology, such as analysis of highly dynamic soluble and membrane-embedded protein complexes or partially ordered protein aggregates, are now within reach. Recent reports of structural studies of G protein–coupled receptors, spliceosomes, and fibrillar specimens illustrate the progress that has been made using cryo-EM methods, and are the main focus of this review.

The sleeping beauty kissed awake: new methods in electron microscopy to study cellular membranes

2017 ◽  
Vol 474 (6) ◽  
pp. 1041-1053 ◽  
Author(s):  
Petr Chlanda ◽  
Jacomine Krijnse Locker
Keyword(s):  
Electron Microscopy ◽  
Cell Biology ◽  
Sleeping Beauty ◽  
Chemical Fixation ◽  
Preparation Methods ◽  
Complementary Method ◽  
The Past ◽  
New Methods ◽  
Technological Advances ◽  
Infected Cells

Electron microscopy (EM) for biological samples, developed in the 1940–1950s, changed our conception about the architecture of eukaryotic cells. It was followed by a period where EM applied to cell biology had seemingly fallen asleep, even though new methods with important implications for modern EM were developed. Among these was the discovery that samples can be preserved by chemical fixation and most importantly by rapid freezing without the formation of crystalline ice, giving birth to the world of cryo-EM. The past 15–20 years are hallmarked by a tremendous interest in EM, driven by important technological advances. Cryo-EM, in particular, is now capable of revealing structures of proteins at a near-atomic resolution owing to improved sample preparation methods, microscopes and cameras. In this review, we focus on the challenges associated with the imaging of membranes by EM and give examples from the field of host–pathogen interactions, in particular of virus-infected cells. Despite the advantages of imaging membranes under native conditions in cryo-EM, conventional EM will remain an important complementary method, in particular if large volumes need to be imaged.

scholarly journals Electron tomography of cells

2011 ◽  
Vol 45 (1) ◽  
pp. 27-56 ◽  
Author(s):  
Lu Gan ◽  
Grant J. Jensen
Keyword(s):  
Electron Microscope ◽  
Cell Biology ◽  
3D Imaging ◽  
Imaging Technique ◽  
Electron Tomography ◽  
Three Dimensional ◽  
Molecular Organization ◽  
Biological Structure ◽  
Structural Cell ◽  
Projection Images

AbstractThe electron microscope has contributed deep insights into biological structure since its invention nearly 80 years ago. Advances in instrumentation and methodology in recent decades have now enabled electron tomography to become the highest resolution three-dimensional (3D) imaging technique available for unique objects such as cells. Cells can be imaged either plastic-embedded or frozen-hydrated. Then the series of projection images are aligned and back-projected to generate a 3D reconstruction or ‘tomogram’. Here, we review how electron tomography has begun to reveal the molecular organization of cells and how the existing and upcoming technologies promise even greater insights into structural cell biology.

scholarly journals Male fertility: review of the publications of January ― March 2020

2020 ◽  
Vol 8 (2) ◽  
pp. 93-98 ◽  
Author(s):  
D. S. Rogozin
Keyword(s):  
Male Infertility ◽  
Selection Criteria ◽  
Assisted Reproductive Technologies ◽  
Seminal Fluid ◽  
Reproductive Technologies ◽  
Practical Significance ◽  
Sperm Dna Fragmentation ◽  
Sperm Dna ◽  
The Impact ◽  
Scimago Journal Rank

The article provides an overview of the most influential papers on the topic of “male infertility”. The selection criteria were the practical significance of the article and the impact factor of the journal in which it was published, according to SCImago Journal Rank (SJR). As a result, we created the list of 10 papers published in the I quarter (January – March) of 2020. The review included articles on the following issues: the effectiveness and safety of antioxidants, zinc and folic acid in the treatment of male infertility; the effect of obesity on sperm DNA-fragmentation; the possibility of obtaining of spermatogonia in children and adolescents with Kleinfelter syndrome; comparison of intracytoplasmic sperm injection (ICSI) and intracytoplasmic morphologically selected spermatozoa injection (IMSI); the reaction of the female reproductive system to the seminal fluid; viral profile in seminal fluid of men receiving the antiretroviral therapy, the effect of sleep duration on fertility and the risk of diabetes mellitus in children, conceived using assisted reproductive technologies.

scholarly journals Structural Cell Biology: Preparing Specimens for Cryo-Electron Tomography Using Focused-Ion-Beam Milling

2014 ◽  
Vol 20 (S3) ◽  
pp. 1222-1223
Author(s):  
Elizabeth Villa ◽  
Miroslava Schaffer ◽  
Ben Engel ◽  
Jürgen Plitzko ◽  
Wolfgang Baumeister
Keyword(s):  
Cell Biology ◽  
Focused Ion Beam ◽  
Electron Tomography ◽  
Ion Beam ◽  
Structural Cell ◽  
Cryo Electron Tomography ◽  
Focused Ion Beam Milling

Cryo-fixation and associated developments in transmission electron microscopy: a cool future for nematology

Nematology ◽  
2016 ◽  
Vol 18 (1) ◽  
pp. 1-14 ◽  
Author(s):  
Wim Bert ◽  
Dieter Slos ◽  
Olivier Leroux ◽  
Myriam Claeys
Keyword(s):  
Electron Microscopy ◽  
Cell Biology ◽  
Electron Tomography ◽  
Light And Electron Microscopy ◽  
Three Dimensional ◽  
Freeze Substitution ◽  
Model Organisms ◽  
Fixation Method ◽  
Dimensional Reconstruction ◽  
Fixation Techniques

At present, the importance of sample preparation equipment for electron microscopy represents the driving force behind major breakthroughs in microscopy and cell biology. In this paper we present an introduction to the most commonly used cryo-fixation techniques, with special attention paid towards high-pressure freezing followed by freeze substitution. Techniques associated with cryo-fixation, such as immunolocalisation, cryo-sectioning, and correlative light and electron microscopy, are also highlighted. For studies that do not require high resolution, high quality results, or the immediate arrest of certain processes, conventional methods will provide answers to many questions. For some applications, such as immunocytochemistry, three-dimensional reconstruction of serial sections or electron tomography, improved preservation of the ultrastructure is required. This review of nematode cryo-fixation highlights that cryo-fixation not only results in a superior preservation of fine structural details, but also underlines the fact that some observations based on results solely obtained through conventional fixation approaches were either incorrect, or otherwise had severe limitations. Although the use of cryo-fixation has hitherto been largely restricted to model organisms, the advantages of cryo-fixation are sufficiently self-evident that we must conclude that the cryo-fixation method is highly likely to become the standard for nematode fixation in the near future.

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