scholarly journals Rapid chemical dehydration of biologic samples for scanning electron microscopy using 2,2-dimethoxypropane.

1977 ◽  
Vol 25 (4) ◽  
pp. 247-251 ◽  
Author(s):  
M D Maser ◽  
J J Trimble
Keyword(s):  
Electron Microscopy ◽  
Scanning Electron Microscopy ◽  
Small Intestine ◽  
Cultured Fibroblasts ◽  
Critical Point Drying ◽  
Mouse Small Intestine ◽  
Fine Surface ◽  
Plastic Embedding ◽  
Rat Trachea ◽  
Scanning Electron

Acidified 2,2-dimethoxypropane (DMP) has been used as a rapid chemical dehydrating agent en route to plastic embedding and ulthrathin sectioning for conventional electron miscroscopy (J Histochem Cytochem 23:107, 1975). We have used DMP to dehydrate biologic specimens prior to critical point drying and metal coating for scanning electron microscopy. There is no difference in either the gross architecture or the fine surface structure of mouse small intestine and trachea, rat trachea and kidney, and cultured fibroblasts, between samples dehydrated in DMP for 5 min to 30 days and those conventionally dehydrated in ethanol or acetone. DMP dehydration is advantageous in speed, economy and apparent completeness.

Preservation of Plant Cell Surfaces For Scanning Electron Microscopy

1973 ◽  
Vol 31 ◽  
pp. 472-473
Author(s):  
Linda M. Sicko ◽  
Thomas E. Jensen
Keyword(s):  
Electron Microscopy ◽  
Scanning Electron Microscopy ◽  
Critical Point ◽  
Filter Paper ◽  
Freeze Drying ◽  
Cell Surfaces ◽  
Air Drying ◽  
Popular Method ◽  
Critical Point Drying ◽  
Scanning Electron

The use of critical point drying is rapidly becoming a popular method of preparing biological samples for scanning electron microscopy. The procedure is rapid, and produces consistent results with a variety of samples. The preservation of surface details is much greater than that of air drying, and the procedure is less complicated than that of freeze drying. This paper will present results comparing conventional air-drying of plant specimens to critical point drying, both of fixed and unfixed material. The preservation of delicate structures which are easily damaged in processing and the use of filter paper as a vehicle for drying will be discussed.

Development of the Foramen Secundum in the Chick as Observed by Scanning Electron Microscopy

1976 ◽  
Vol 34 ◽  
pp. 212-213
Author(s):  
M.J.C. Hendrix ◽  
D.E. Morse
Keyword(s):  
Electron Microscopy ◽  
Scanning Electron Microscopy ◽  
Procaine Hydrochloride ◽  
Cardiac Anomaly ◽  
Chick Embryos ◽  
Congenital Cardiac Anomaly ◽  
Septal Defects ◽  
Critical Point Drying ◽  
Scanning Electron

Atrial septal defects are considered the most common congenital cardiac anomaly occurring in humans. In studying the normal sequential development of the atrial septum, chick embryos of the White Leghorn strain were prepared for scanning electron microscopy and the results were then extrapolated to the human heart. One-hundred-eighty chick embryos from 2 to 21 days of age were removed from their shells and immersed in cold cacodylate-buffered aldehyde fixative . Twenty-four embryos through the first week post-hatching were perfused in vivo using cold cacodylate-buffered aldehyde fixative with procaine hydrochloride. The hearts were immediately dissected free and remained in the fixative a minimum of 2 hours. In most cases, the lateral atrial walls were removed during this period. The tissues were then dehydrated using a series of ascending grades of ethanol; final dehydration of the tissues was achieved via the critical point drying method followed by sputter-coating with goldpalladium.

Intracellular structures of rapid frozen biological samples observed by high-resolution low-temperature Scanning Electron Microscopy

1989 ◽  
Vol 47 ◽  
pp. 736-737
Author(s):  
T. Inoué ◽  
H. Koike
Keyword(s):  
Electron Microscopy ◽  
Scanning Electron Microscopy ◽  
Low Temperature ◽  
Liquid Nitrogen ◽  
Specimen Preparation ◽  
Chemical Fixation ◽  
Brown Adipose ◽  
Critical Point Drying ◽  
Temperature Scanning ◽  
Scanning Electron

Low temperature scanning electron microscopy (LTSEM) is useful to avoid artifacts such as deformation and extraction, because specimens are not subjected to chemical fixation, dehydration and critical-point drying. Since Echlin et al developed a LTSEM, many techniques and instruments have been reported for observing frozen materials. However, intracellular structures such as mitochondria and endoplasmic reticulum have been unobservable by the method because of the low resolving power and inadequate specimen preparation methods. Recently, we developed a low temperature SEM that attained high resolutions. In this study, we introduce highly magnified images obtained by the newly developed LTSEM, especially intracellular structures which have been rapidly frozen without chemical fixation.[Specimen preparations] Mouse pancreas and brown adipose tissues (BAT) were used as materials. After the tissues were removed and cut into small pieces, the specimen was placed on a cryo-tip and rapidly frozen in liquid propane using a rapid freezing apparatus (Eiko Engineering Co. Ltd., Japan). After the tips were mounted on the specimen stage of a precooled cryo-holder, the surface of the specimen was manually fractured by a razor blade in liquid nitrogen. The cryo-holder was then inserted into the specimen chamber of the SEM (ISI DS-130), and specimens were observed at the accelerating voltages of 5-8 kV. At first the surface was slightly covered with frost, but intracellular structures were gradually revealed as the frost began to sublimate. Gold was then coated on the specimen surface while tilting the holder at 45-90°. The holder was connected to a liquid nitrogen reservoir by means of a copper braid to maintain low temperature.

Criteria for the evaluation of low-temperature Scanning Electron Microscopy

1986 ◽  
Vol 44 ◽  
pp. 902-903
Author(s):  
Alan Beckett
Keyword(s):  
Electron Microscopy ◽  
Scanning Electron Microscopy ◽  
Low Temperature ◽  
Conformational Changes ◽  
Chemical Vapour ◽  
List Type ◽  
Dimensional Changes ◽  
Critical Point Drying ◽  
Temperature Scanning ◽  
Scanning Electron

Low temperature scanning electron microscopy (LTSEM) has been evaluated with special reference to its application to the study of morphology and development in microorganisms. A number of criteria have been considered and have proved valuable in assessing the standard of results achieved. To further aid our understanding of these results, it has been necessary to compare those obtained by LTSEM with those from more conventional preparatory procedures such as 1) chemical fixation, dehydration and critical point-drying; 2) freeze-drying with or without chemical vapour fixation before hand.The criteria used for assessing LTSEM for the above purposes are as follows: 1)Specimen immobilization and stabilization2)General preservation of external morphology3)General preservation of internal morphology4)Exposure to solvents5)Overall dimensional changes6)Cell surface texture7)Differential conformational changes8)Etching frozen-hydrated material9)Beam damage10)Specimen resolution11)Specimen life

Vergleich verschiedener Immun-Dekorationsmethoden für die Rasterelektronenmikroskopie am Beispiel eines Protein-Antigens auf der Oberfläche der Hefe Candida albicans/ A Comparison of Decoration Techniques for the Demonstration of Immunocomplexes by Scanning Electron Microscopy: Labeling of a Protein Antigen on the Surface of the Yeast Candida albicans

1985 ◽  
Vol 40 (7-8) ◽  
pp. 539-550 ◽  
Author(s):  
Margarete Borg
Keyword(s):  
Electron Microscopy ◽  
Scanning Electron Microscopy ◽  
Candida Albicans ◽  
Protein A ◽  
Peritoneal Macrophages ◽  
Polystyrene Latex ◽  
Target Antigen ◽  
Protein Antigens ◽  
Critical Point Drying ◽  
Scanning Electron

Abstract The labeling of immunocomplexes for scanning electron microscopy (SEM) is a fairly new technique, and the various procedures, that have been proposed, have not yet been compared. Such comparative evaluation was performed with Candida protease as a target antigen. This secretory enzyme of the opportunistic yeast Candida albicans can be localized on the surface of fungal blastopores and mycelia, both after growth in proteinaceous medium and upon infection of murine peritoneal macrophages. The presence of the protease antigen was confirmed by immunofluorescence and by immunoperoxidase-light microscopy. The decoration of protease - anti protease complexes for SEM was attempted with colloids derived from the immunoperoxidase reaction, by the immunogold technique, and by antibodies linked to beads of synthetic polymers (polystyrene, polymethacrylate, polyacrolein). In addition, inactivated Staphylococcus aureus was used, which binds to antibodies through its protein-A. The high resolution by SEM of surface structures was matched only by the colloid based decoration techniques. All conjugates with beads suffered from inconsistent binding, which did not correspond with the distribution of the surface antigen. The comparatively best result with beads was obtained with polystyrene (Latex). Colloid based techniques in addition allow for critical point drying, which cannot be applied to synthetic beads in the usual manner.

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