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The Feulgen reaction after hydrolysis at room temperature 总被引:2,自引:0,他引:2
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Tosisuke Hiraoka 《Histochemistry and cell biology》1973,35(4):283-296
Summary Electrophoretic means of separation revealed the presence of as many as five reaction products in Schiff-apurinic acid reaction at the maximum. They differed not only in their absorption maxima, but also in their ratios of apurinic acid phosphorus to fuchsin moiety. Some considerations on the reaction mechanism to account for the occurrence of these multiple reaction products have been made. The stoichiometry of Schiff-apurinic acid reaction was studied with respect to the main product responsible for the presentation of reaction color. A reaction product consisting of six or eight atom moles of apurinic acid phosphorus per mole of fuchsin moiety is to be formed, provided that the reagent of infinite concentration is used. From theoretical view point, a reaction product consisting of four atom moles of apurinic acid phosphorus per mole of fuchsin moiety is to be expected with the reagent of infinite concentration, provided that apurinic acid retains essentially the nucleotide sequence of its parent desoxyribonucleic acid except for some modification of the original purin nucleotide groups to react as aldehyde moieties, and provided that the reaction proceeds at a constant rate irrespective of the concentrations of the reagent. 相似文献
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Wiebke Schlörmann Frank Steiniger Walter Richter Roland Kaufmann Gerd Hause Cornelius Lemke Martin Westermann 《Histochemistry and cell biology》2010,133(2):223-228
Caveolae were defined as flask- or omega-shaped plasma membrane invaginations, abundant in adipocytes, fibroblasts, endothelial
and smooth muscle cells. The major protein component of caveolar membranes is an integral membrane protein named caveolin.
We compared the freeze-fracture behavior of caveolae in glutaraldehyde-fixed and cryofixed mouse fibroblast cells and found
distinct differences. In glutaraldehyde-fixed cells almost all caveolae were cross-fractured through their pore and only very
few caveolar membranes were membrane-fractured. We found the reverse situation in rapid frozen cells without any chemical
fixation where most of the caveolae were membrane-fractured, showing different degrees of invagination from nearly flat to
deeply invaginated. In ultrathin sections of glutaraldehyde-fixed heart endothelial cells, caveolae exhibit the well known
omega-like shape. In high-pressure frozen, freeze-substituted and low temperature embedded heart endothelial cells, the caveolae
frequently exhibit a cup-like shape without any constriction or pore. The cup-like caveolar shape could also be shown by tilt
series analysis of freeze-fracture replicas obtained from cryofixed cells. Freeze-fracture immunolabeling of caveolin-1 revealed
a lateral belt-like caveolin alignment. These findings point out that the constricted “neck” region of caveolae in most cases
is an effect that is caused and intensified by the glutaraldehyde fixation. Our data indicate that caveolae in vivo show all
degrees of invagination from nearly flat via cup-like depressed to in a few cases omega-like. 相似文献
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The Feulgen reaction 75 years on 总被引:9,自引:0,他引:9
The Feulgen reaction proposed by Feulgen and Rossenbeck 75 years ago is one of the cytohistochemical reactions most widely
used in biology and medicine. It allows DNA in situ to be specifically stained based on the reaction of Schiff or Schiff-like
reagents with aldehyde groups engendered in the deoxyribose molecules by HCl hydrolysis. The staining intensity is proportional
to the DNA concentration. Current applications of the Feulgen reaction are mainly concerned with DNA quantification in cell
nuclei by image cytometry for ploidy evaluation in tumor pathology. From the morphological point of view, specific demonstration
of DNA in cell structures at the light microscopic level is very little used nowadays. On the other hand, application of the
Feulgen principles to electron microscopy have recently allowed specific DNA-staining procedures to be developed for the study
of the structural organization of DNA in situ.
Accepted: 13 January 1999 相似文献
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Methyl green-pyronin staining has been used for localization of RNA and DNA in chick and mouse embryonic tissues and in insect larval salivary glands. Glutaraldehyde or tricholoracetic acid-lanthanum acetate (TCA-LA) was used as fixative and paraffin wax or Araldite was used as embedding medium. For good results the following are specially desirable: fixation with 2.5% glutaraldehyde, dehydration in alcohols for short time, and the use of fresh staining solutions. After TCA-LA fixation the final results are much less specific. The digestion with RNAse appears essential for the detection of RNA because pyronin does not seem to be entirely specific to RNA. The results show that glutaraldehyde a common fixative for electron microscopic work, is particularly suitable fixative for light microscopic cytochemical investigations if followed by methyl green-pyronin staining; furthermore, methyl green-pyronin staining after glutaraldehyde fixation can be carried out on Araldite sections. 相似文献
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STEDMAN E 《The Biochemical journal》1950,47(4):508-512
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Autoradiographs were prepared of sections of the ovary of Dytiscus marginalis labelled with thymidine-3H after each successive step of the Feulgen reaction and after treatment with each separate component of the Schiff's reagent. Results of grain counts over ovarian nurse cells showed that losses of thymidine-3H activity occur not only during hydrolysis but also during the successive steps of the Feulgen reaction. It is suggested that the latter decrease in radioactivity may depend on the extraction of fragments of apurinic acid from the sections. An emulsion desensitizing effect has also been observed in sections stained with basic fuchsin alone; this effect appears, however, to be strongly counteracted by the metabisulphite present in the Schiff's reagent. 相似文献
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New glutaraldehyde fixation procedures 总被引:1,自引:0,他引:1
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A technique is described for performing the Feulgen reaction for DNA on cells and tissues fixed in glutaraldehyde. Blockade free aldehydes by reducing them with fresh 0.5% NaBH4 in 1% NaH2PO4 for 1 hr at room temperature, then rinse in water. Follow by a Feulgen reaction (hydrolysis at room temperature in 6 N HCI for 20 min, Schiff's reagent for 60 min.). Controls assure the completeness and irreversibility of the borohydride blockade. Cytophotometry shows that the DNA content per nucleus is unaffected by the blockade procedure. 相似文献
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The reaction of glutaraldehyde with tissue lipids 总被引:5,自引:0,他引:5
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Locust neural lamella and Calpodes connective tissue fixed in glutaraldehyde have a fibrous component which stains after reaction with DAB and osmication and after staining sections with PTA. The fibers also stain when fixed in glutaraldehyde with tannic acid followed by osmication and section staining with lead citrate. 相似文献
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