Rapid-freeze fixation and substitution improves tissue preservation of microspores and tapetum in Brassica napus

2000 ◽  
Vol 12 (4) ◽  
pp. 237-240 ◽  
Author(s):  
J. H. E. Ross ◽  
C. Milanesi ◽  
D. J. Murphy ◽  
Mauro Cresti
Keyword(s):  
Brassica Napus ◽  
Tissue Preservation ◽  
Rapid Freeze Fixation ◽  
Freeze Fixation

Ultrastructural study of archaeological Vitis vinifera L. seeds using rapid-freeze fixation and substitution

2009 ◽  
Vol 41 (6) ◽  
pp. 443-447 ◽  
Author(s):  
Claudio Milanesi ◽  
Rita Vignani ◽  
Andrea Ciacci ◽  
Alessandra Nardini ◽  
Marco Valenti ◽  
...  
Keyword(s):  
Vitis Vinifera ◽  
Ultrastructural Study ◽  
Vitis Vinifera L ◽  
Rapid Freeze Fixation ◽  
Freeze Fixation

Ultrastructure of scales in a teleost (Carassius auratus L.) after use of rapid freeze-fixation and freeze-substitution

1982 ◽  
Vol 223 (2) ◽  
pp. 349-367 ◽  
Author(s):  
Louise Zylberberg ◽  
G. Nicolas
Keyword(s):  
Carassius Auratus ◽  
Freeze Substitution ◽  
Rapid Freeze Fixation ◽  
Freeze Fixation

A method for rapid freeze fixation of plant cells

PROTOPLASMA ◽  
1986 ◽  
Vol 131 (2) ◽  
pp. 153-165 ◽  
Author(s):  
Susan A. Lancelle ◽  
D. A. Callaham ◽  
P. K. Hepler
Keyword(s):  
Plant Cells ◽  
Rapid Freeze Fixation ◽  
Freeze Fixation

Epoxy Resin Embedment

1968 ◽  
Vol 26 ◽  
pp. 100-101
Author(s):  
J. G. Adams ◽  
M. M. Campbell ◽  
H. Thomas ◽  
J. J. Ghldonl
Keyword(s):  
Electron Microscopy ◽  
Electron Beam ◽  
Epoxy Resin ◽  
Light Microscope ◽  
Epoxy Resins ◽  
Image Contrast ◽  
Tissue Preservation ◽  
Resin System ◽  
Excellent Quality

Since the introduction of epoxy resins as embedding material for electron microscopy, the list of new formulations and variations of widely accepted mixtures has grown rapidly. Described here is a resin system utilizing Maraglas 655, Dow D.E.R. 732, DDSA, and BDMA, which is a variation of the mixtures of Lockwood and Erlandson. In the development of the mixture, the Maraglas and the Dow resins were tested in 3 different volumetric proportions, 6:4, 7:3, and 8:2. Cutting qualities and characteristics of stability in the electron beam and image contrast were evaluated for these epoxy mixtures with anhydride (DDSA) to epoxy ratios of 0.4, 0.55, and 0.7. Each mixture was polymerized overnight at 60°C with 2% and 3% BDMA.Although the differences among the test resins were slight in terms of cutting ease, general tissue preservation, and stability in the beam, the 7:3 Maraglas to D.E.R. 732 ratio at an anhydride to epoxy ratio of 0.55 polymerized with 3% BDMA proved to be most consistent. The resulting plastic is relatively hard and somewhat brittle which necessitates trimming and facing the block slowly and cautiously to avoid chipping. Sections up to about 2 microns in thickness can be cut and stained with any of several light microscope stains and excellent quality light photomicrographs can be taken of such sections (Fig. 1).

Immunocytochemistry after fast freeze fixation-freeze substitution: Advantages and limits

1990 ◽  
Vol 48 (3) ◽  
pp. 42-43
Author(s):  
Marie-Thérèse Nicolas
Keyword(s):  
Osmium Tetroxide ◽  
Freeze Substitution ◽  
Acrylic Resin ◽  
Expected Improvement ◽  
List Type ◽  
Chemical Fixation ◽  
Freeze Fixation ◽  
Liquid Chemical ◽  
Luciferase Enzyme ◽  
Lower Background

An alternative to aqueous chemical fixation consists in immobilizing physically the specimen by freezing it as fast as possible without using any cryoprotectant. This Fast Freeze Fixation (FFF) followed by Freeze Substitution (FS) avoids osmotic artefacts due to the slow penetration of liquid chemical fixative. Associated with Immuno-Gold labeling (IGS), FFF-FS allows a more precise localization of antigens.Using the bioluminescent bacteria Vibrio harveyi, a comparison of IGS with an antibody directed against its luciferase (enzyme of the luminescent reaction) has been done after liquid chemical fixation versus FFFFS. This later technique, beside an expected improvement of the ultrastructure always shows a better preservation of antigenicity and a lower background. In the case of FFF-FS technique (Figure 3):–labeling in acrylic resin (LRWhite) is 2 to 4 fold more intense than in epoxy resin (Epon),–but the ultrastructure is always better in Epon.–but the ultrastructure is always better in Epon.–The addition of fixatives in the substitution medium, results in a decrease of labeling which is more important in the case of a mixture of fixatives than with osmium tetroxide alone; with one exception: the substitution with glutaraldehyde which produces a dramatic increase in the density of the labeling but also, at the same time, a swelling of the cells of about 30%.

Osmolarity and Artificial Cell Damage in Ischemic Myocardium

1990 ◽  
Vol 48 (3) ◽  
pp. 262-263
Author(s):  
Richard Montione ◽  
Muhammad Ashraf
Keyword(s):  
Cell Damage ◽  
Uranyl Acetate ◽  
Tissue Preservation ◽  
Rat Myocardium ◽  
En Bloc ◽  
Cellular Changes ◽  
Artificial Cell ◽  
Cacodylate Buffer ◽  
Isotonic Buffer ◽  
Perfusion Fixation

Osmolarity of a fixative vehicle has long been known to have an effect on the tissue preservation. An increase in tissue osmolarity occurs in ischemia-damaged tissue and affects the morphology. In this study, we examined cellular changes in ischemic rat myocardium induced by varying fixative toxicity.Rats were sacrificed by decapitation and the hearts immediately removed and retrogradily perfused through the aorta with anoxic Kurbs-Henseleit medium. Hearts were then placed in a bag with a small amount of medium at 37°C for 90 minutes. Hearts were perfusion-fixed using 2% glutaraldehyde in 0.1 M cacodylate buffer pH -7.3 at three osmolarities. The isotonic buffer was adjusted to 311 mOsm/kg using D-manitol. Hypertonic buffers were adjusted to 375 and 400 mOsm/kg. One-half hour after perfusion fixation, the hearts were sliced and cut into small blocks and allowed to fix overnight at 4°C. Blocks were post fixed in osmium, en bloc stained in uranyl acetate, dehydrated in ethanol and embedded in Spurr medium.

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