Tissue printing for scanning electron microscopy and microanalysis

1993 ◽  
Vol 51 ◽  
pp. 140-141
Author(s):  
Barbara A. Reine
Keyword(s):  
Critical Point ◽  
Chemical Constituents ◽  
Plant Cells ◽  
Plant Morphology ◽  
Specimen Preparation ◽  
Normal Surface ◽  
Tissue Printing ◽  
Critical Point Drying ◽  
Scanning Electron

The study of plant morphology and plant cells in the scanning electron microscope is often compromised by the limitations of specimen preparation techniques. Simple natural dehydration usually results in unacceptable shrinkage and distortion of the normal surface morphology of plant cells. Chemical fixation followed by critical point drying or some substitute for critical point drying such as Peldri II or HMDS (hexamethyldisilazane) improves morphological results but still imparts artifacts, adds chemical constituents to the specimen, and requires the use of toxic chemicals, a hood, and much time.One technique that eliminates many of these disadvantages and is even suitable for specimen preparation in the field is tissue printing. For low magnification imaging and chemical analysis its “elegant simplicity” (2) is compelling. Historically, the application of tissue printing has been in connection with optical microscopy (1,2). However, this technique works very well for low magnification SEM and associated elemental characterization of residues by x-ray microanalysis.

scholarly journals Tissue Printing for Scanning Electron Microscopy and Microanalysis

1994 ◽  
Vol 2 (1) ◽  
pp. 8-8
Author(s):  
Barbara A. Reine
Keyword(s):  
Optical Microscopy ◽  
Chemical Constituents ◽  
Plant Cells ◽  
Plant Morphology ◽  
Specimen Preparation ◽  
Normal Surface ◽  
Stem Anatomy ◽  
Tissue Printing ◽  
Critical Point Drying ◽  
Scanning Electron

The study of plant morphology and plant cells in the scanning electron microscope is often compromised by the limitations of specimen preparation techniques. Air drying usually results in unacceptable shrinkage and distortion of the normal surface morphology of plant cells. Chemical fixation followed by critical point drying or chemical drying using fluorocarbon compounds improves morphological results but still imparts artifacts, adds chemical constituents to the specimen, and requires the use of toxic chemicals, a hood, and much time.One technique that eliminates many of these disadvantages and is even suitable for specimen preparation in the field is tissue printing. For easy, quick recording of stem anatomy and collection of cell exudates for subsequent analysis, its “elegant simlicity” is compelling. Historically, the application of tissue printing has been in connection with optical microscopy. However, this technique works very well for SEM and associated elemental characterization of residues by x-ray microanalysis. The tissue print technique applied to SEM is much the same as for optical microscopy.

The Critical Point Drying Method Applied to Scanning Electron Microscopy of L-929 Cells

1970 ◽  
Vol 28 ◽  
pp. 278-279
Author(s):  
Charles TurnbiLL ◽  
Delbert E. Philpott
Keyword(s):  
Electron Microscopy ◽  
Carbon Dioxide ◽  
Surface Tension ◽  
Critical Point ◽  
Freeze Drying ◽  
Attractive Alternative ◽  
Crystal Formation ◽  
Critical Point Drying ◽  
Time Required ◽  
Scanning Electron

The advent of the scanning electron microscope (SCEM) has renewed interest in preparing specimens by avoiding the forces of surface tension. The present method of freeze drying by Boyde and Barger (1969) and Small and Marszalek (1969) does prevent surface tension but ice crystal formation and time required for pumping out the specimen to dryness has discouraged us. We believe an attractive alternative to freeze drying is the critical point method originated by Anderson (1951; for electron microscopy. He avoided surface tension effects during drying by first exchanging the specimen water with alcohol, amy L acetate and then with carbon dioxide. He then selected a specific temperature (36.5°C) and pressure (72 Atm.) at which carbon dioxide would pass from the liquid to the gaseous phase without the effect of surface tension This combination of temperature and, pressure is known as the "critical point" of the Liquid.

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.

Modified gach technique vs critical point drying: Shrinkage analysis for SEM

1987 ◽  
Vol 45 ◽  
pp. 954-955
Author(s):  
B. Thompson ◽  
N. Sculov ◽  
R.E. Crang
Keyword(s):  
Critical Point ◽  
Calf Serum ◽  
Drying Shrinkage ◽  
Specimen Preparation ◽  
Cell Shrinkage ◽  
Whole Cells ◽  
Critical Point Drying ◽  
Prepared Material ◽  
Inverted Microscope ◽  
Phosphate Buffered Saline

The use of co-polymerized glutaraldehyde-carbohydrazide (GACH) was proposed for specimen preparation in scanning electron microscopy (SEM) as a means of avoiding dehydration in organic solvents, and to provide dimensionally stable biological specimens through a process of air-drying. It has been assumed that shrinkage of specimens prepared by the GACH technique should be less than that of conventionally-prepared material by critical point drying (CPD). In a previous study, Bell has reported significant shrinkage of whole cells for SEM. This report compares cell shrinkage in GACH and CPD preparations.Fibroblasts from newborn rats were grown on collagen-coated glass cover-slips (with alpha numeric grids etched onto the surface of the coverslips) in Eagle's minimum essential medium + 10% fetal calf serum for 7 d. (3). Using an inverted microscope with phase-contrast optics, micrographs were taken of the cultures in their live state and 1 h. after fixation with 2.5% glutaraldehyde in Dulbecco's phosphate buffered saline (Figs. 1 and 3).

Scanning electron microscopy of plant cells using a variable-pressure SEM and cryogenic techniques

1993 ◽  
Vol 51 ◽  
pp. 260-261
Author(s):  
M. Yamada ◽  
K. Ueda ◽  
K. Kuboki ◽  
H. Matsushima ◽  
S. Joens
Keyword(s):  
Electron Microscopy ◽  
Scanning Electron Microscopy ◽  
Saturated Vapor ◽  
Plant Cells ◽  
Variable Pressure ◽  
Specimen Preparation ◽  
Operating Pressure ◽  
Vapor Pressures ◽  
Low Temperature Microscopy ◽  
Scanning Electron

Use of variable Pressure SEMs is spreading among electron microscopists The variable Pressure SEM does not necessarily require specimen Preparation such as fixation, dehydration, coating, etc which have been required for conventional scanning electron microscopy. The variable Pressure SEM allows operating Pressure of 1˜270 Pa in specimen chamber It does not allow microscopy of water-containing specimens under a saturated vapor Pressure of water. Therefore, it may cause shrink or deformation of water-containing soft specimens such as plant cells due to evaporation of water. A solution to this Problem is to lower the specimen temperature and maintain saturated vapor Pressures of water at low as shown in Fig. 1 On this technique, there is a Published report of experiment to have sufficient signal to noise ratio for scondary electron imaging at a relatively long working distance using an environmental SEM. We report here a new low temperature microscopy of soft Plant cells using a variable Pressure SEM (Hitachi S-225ON).

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.

Pulmonary microcirculation: tubules rather than sheet and post

1982 ◽  
Vol 53 (2) ◽  
pp. 510-515 ◽  
Author(s):  
W. G. Guntheroth ◽  
D. L. Luchtel ◽  
I. Kawabori
Keyword(s):  
Electron Microscope ◽  
Scanning Electron Microscope ◽  
Critical Point ◽  
Ringer Solution ◽  
Basic Component ◽  
Pulmonary Vasculature ◽  
Critical Point Drying ◽  
Pulmonary Microcirculation ◽  
Scanning Electron

We examined latex casts of the pulmonary microcirculation with the scanning electron microscope (SEM). Mature rats were anesthetized and ventilated; the pulmonary vasculature was washed out with lactated Ringer solution and then filled with a mixture of Geon latexes. The airways were filled with glutaraldehyde with resulting transmural vascular pressures of 10 cmH2O. After critical-point drying and corrosive removal of the lung tissue, SEM studies of the vascular replicas revealed two distinct patterns of pulmonary microcirculation: 1) sparse, long, tubular capillaries that comprise the thin subpleural layer and appear as “filler” in the peribronchial spaces; and 2) alveolar microcirculation that is composed of tightly matted, intersecting tubules, shorter but of the same diameter as type 1, in spherical array in two layers. The alveolar capillaries at low magnification appear superficially as sheets; however, the detailed morphology is not consistent with the sheet-and-post model. We conclude that the basic component of the pulmonary microcirculation is tubular and not different from other capillary beds except in density.

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