Pre-embedding Method of Electron Microscopy for Glycan Localization in Mammalian Tissues and Cells Using Lectin Probes

2016 ◽  
pp. 259-267 ◽  
Author(s):  
Yoshihiro Akimoto ◽  
Kuniaki Takata ◽  
Hayato Kawakami
Keyword(s):  
Electron Microscopy ◽  
Embedding Method ◽  
Mammalian Tissues

Rapid Chemical Dehydration of Mammalian Tissues for Scanning Electron Microscopy using 2 ,2-Dimethoxypropane

1976 ◽  
Vol 34 ◽  
pp. 340-341
Author(s):  
Morton D Maser ◽  
John J. Trimble
Keyword(s):  
Electron Microscopy ◽  
Scanning Electron Microscopy ◽  
Transmission Electron Microscopy ◽  
Laboratory Mouse ◽  
Transmission Electron ◽  
Critical Point Drying ◽  
Mammalian Tissues ◽  
Cervical Dislocation ◽  
Structural Preservation ◽  
Scanning Electron

Muller and Jacks used 2,2-dimethoxypropane (DMP) as a dehydrating agent for biological specimens prior to embedment, ultrathin sectioning, and examination by transmission electron microscopy. Acidified DMP reacts endothermically with water to produce methanol and acetone. Muller and Jacks demonstrated excellent fine structural preservation in a variety of specimens dehydrated with DMP.We have used the same method, prior to critical point drying, to prepare specimens for scanning electron microscopy (SEM) . Portions of the trachea, small intestine, and kidney from a laboratory mouse killed by cervical dislocation were fixed for 16 hours in cold 3% glutaraldehyde in 0.1 M phosphate buffer, pH 7.2.

Water Containing Epon 812/815 Embedding Method for Electron Microscopy

1980 ◽  
Vol 38 ◽  
pp. 642-643
Author(s):  
Yoshiya Shinagawa ◽  
Yasuko Shlnagawa ◽  
Sadao Uchida
Keyword(s):  
Electron Microscopy ◽  
Epoxy Resins ◽  
Embedding Method ◽  
Thin Sectioning

The special types of epoxy resins such as Durcupan or Quetol 651 have been used as water-miscible embedding media for electron microscopy hitherto. However, difficulties exist in handling of these resins, especially in thin-sectioning. We have developed a method of polymerization of the conventional epoxy resins, Epon 812, 815 (Shell Co.) and Luveak 812 (Nakarai Co.) in the presence of water. The authorsl) previously reported melamine resins as water-containing embedding media which have been recently sold by Nakarai Co. as Luveak A and B. The melamine resins as well as aldehyde or urea resins have atypical electron staining. The water-containing epoxy resins embedding method in this report provides usual electron staining and facile sectioning like as the traditional Epon embedding method. Negatively stained micrograph in part is obtained in this method (Fig. la).

A Quick Embedding Method for Light and Electron Microscopy of Textile Fibers

1991 ◽  
Vol 66 (3) ◽  
pp. 122-124 ◽  
Author(s):  
E. K. Boylston ◽  
O. Hinojosa ◽  
J. J. Hebert
Keyword(s):  
Electron Microscopy ◽  
Light And Electron Microscopy ◽  
Embedding Method ◽  
Textile Fibers

scholarly journals A flat embedding method for transmission electron microscopy reveals an unknown mechanism of tetracycline

2021 ◽  
Vol 4 (1) ◽  
Author(s):  
Michaela Wenzel ◽  
Marien P. Dekker ◽  
Biwen Wang ◽  
Maroeska J. Burggraaf ◽  
Wilbert Bitter ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Transmission Electron Microscopy ◽  
Large Scale ◽  
Embedding Method ◽  
Peripheral Membrane Proteins ◽  
Transmission Electron ◽  
Cell Sample ◽  
Flat Embedding ◽  
Induced Changes ◽  
Membrane Deformations

AbstractTransmission electron microscopy of cell sample sections is a popular technique in microbiology. Currently, ultrathin sectioning is done on resin-embedded cell pellets, which consumes milli- to deciliters of culture and results in sections of randomly orientated cells. This is problematic for rod-shaped bacteria and often precludes large-scale quantification of morphological phenotypes due to the lack of sufficient numbers of longitudinally cut cells. Here we report a flat embedding method that enables observation of thousands of longitudinally cut cells per single section and only requires microliter culture volumes. We successfully applied this technique to Bacillus subtilis, Escherichia coli, Mycobacterium bovis, and Acholeplasma laidlawii. To assess the potential of the technique to quantify morphological phenotypes, we monitored antibiotic-induced changes in B. subtilis cells. Surprisingly, we found that the ribosome inhibitor tetracycline causes membrane deformations. Further investigations showed that tetracycline disturbs membrane organization and localization of the peripheral membrane proteins MinD, MinC, and MreB. These observations are not the result of ribosome inhibition but constitute a secondary antibacterial activity of tetracycline that so far has defied discovery.

scholarly journals New flat embedding method for transmission electron microscopy reveals an unknown mechanism of tetracycline

2019 ◽  
Author(s):  
Michaela Wenzel ◽  
Marien P. Dekker ◽  
Biwen Wang ◽  
Maroeska J. Burggraaf ◽  
Wilbert Bitter ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Transmission Electron Microscopy ◽  
Cell Membrane ◽  
Large Scale ◽  
Public Health Problem ◽  
Fixation Method ◽  
Bacterial Cells ◽  
Embedding Method ◽  
Transmission Electron ◽  
Flat Embedding

AbstractTransmission electron microscopy (TEM) is an important imaging technique in bacterial research and requires ultrathin sectioning of resin embedding of cell pellets. This method consumes milli- to deciliters of culture and results in sections of randomly orientated cells. For rod-shaped bacteria, this makes it exceedingly difficult to find longitudinally cut cells, which precludes large-scale quantification of morphological phenotypes. Here, we describe a new fixation method using either thin agarose layers or carbon-coated glass surfaces that enables flat embedding of bacteria. This technique allows for the observation of thousands of longitudinally cut rod-shaped cells per single section and requires only microliter culture volumes. We successfully applied this technique to Gram-positive Bacillus subtilis, Gram-negative Escherichia coli, the tuberculosis vaccine strain Mycobacterium bovis BCG, and the cell wall-lacking mycoplasma Acholeplasma laidlawii. To assess the potential of the technique to quantify morphological phenotypes, we examined cellular changes induced by a panel of different antibiotics. Surprisingly, we found that the ribosome inhibitor tetracycline causes significant deformations of the cell membrane. Further investigations showed that the presence of tetracycline in the cell membrane changes membrane organization and affects the peripheral membrane proteins MinD, MinC, and MreB, which are important for regulation of cell division and elongation. Importantly, we could show that this effect is not the result of ribosome inhibition but is a secondary antibacterial activity of tetracycline that has defied discovery for more than 50 years.SignificanceBacterial antibiotic resistance is a serious public health problem and novel antibiotics are urgently needed. Before a new antibiotic can be brought to the clinic, its antibacterial mechanism needs to be elucidated. Transmission electron microscopy is an important tool to investigate these mechanisms. We developed a flat embedding method that enables examination of many more bacterial cells than classical protocols, enabling large-scale quantification of phenotypic changes. Flat embedding can be adapted to most growth conditions and microbial species and can be employed in a wide variety of microbiological research fields. Using this technique, we show that even well-established antibiotics like tetracycline can have unknown additional antibacterial activities, demonstrating how flat embedding can contribute to finding new antibiotic mechanisms.

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