Effect of osmotic stress on thylakoid fine structure inPorphyra umbilicalis

Summary The fine structural organization of thylakoid membranes in intact cells ofPorphyra umbilicalis, an intertidal red alga, was studied using the freeze-fracture method with special emphasis on changes induced by hypo- and hyperosmotic stresses. In osmotically adapted plants the density of intramembraneous particles on the PF-face increases considerably in the osmotic range from 5-fold diluted to 6-fold concentrated artifical seawater medium ASP₁₂, while that on the EF-face remains constant. The size of the particles on both fracture faces decreases strongly from extreme hypoosmotic to extreme hyperosmotic stress. These findings are discussed with relation to their biological significance.The author is member of the Arbeitsgemeinschaft für Elektronenmikroskopie an der Tierärztlichen Hochschule Hannover.     

Studien über die endokrinen Zellen des Magendarmtraktes

Zusammenfassung In Gangepithelien, Drüsenacini und Langerhansschen Inseln des Pankreas der Katze werden EC-Zellen elektronenmikroskopisch nachgewiesen. In fluoreszenzmikroskopischen Untersuchungen finden sich nach Formaldehydbedampfung stark gelb fluoreszierende, serotoninhaltige Zellen, die nach dem gleichen histotopographischen Muster verteilt sind. Diese und vergleichende Untersuchungen am Pankreas der Ratte ergaben, daß die EC-Zelle der Elektronenmikroskopie der serotoninhaltigen Zelle der Fluoreszenzmikroskopie entspricht. Im Analogschluß werden die Ergebnisse auf die EC-Zellen im Magendarmepithel übertragen. Die Abgrenzung der EC-Zelle von anderen endokrinen Zellen des Magendarmtrakts und die Rolle der EC-Zelle im Serotoninstoffwechsel werden diskutiert.

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Studies on the endocrine cells of the gastrointestinal tractI. Histotopographic, fluorescence microscopic and ultrastructural investigations of the cat and rat

Summary By means of electron microscopy EC-cells were found in the epithelium of the ducts, in the acini and in the islets of Langerhans of the feline pancreas. Investigations carried out with the method of formol induced fluorescence showed yellow strongly fluorescent serotonin-containing cells following the same distribution pattern. These results and concomitant research on the pancreas of the rat lead to the conclusion that the EC-cell, seen ultrastructurally, corresponds to the serotonin-containing cell of fluorescence microscopy. The results are applied to the EC-cells of the gastrointestinal epithelium. The difference between the EC-cell and the other endocrine cells of the gastro-intestinal tract and the role of EC-cells in serotonin-metabolism are discussed.

Mit Unterstützung durch die Deutsche Forschungsgemeinschaft, Antrag Fo 77, 1–4.     

Zur histochemischen Ionenlokalisation mit Hilfe der Elektronenmikroskopie unter besonderer Berücksichtigung der Chloridreaktion

Zusammenfassung Nach einer Erörterung der Voraussetzungen, Methodik und Grenzen der histochemischen Ionenlokalisation mit Hilfe der Elektronenmikroskopie werden an einigen Beispielen der Chloridfällung Fragen der Interpretation und Diffusionsartefakte behandelt. Durch Kontrollversuche an der Niere, sowie an Beleg- und Chloridzellen wird die Spezifität der histochemischen Chloridmethode aufgezeigt und das Reaktionsprodukt im Falle der Chloridzellen durch Elektronenbeugung einwandfrei identifiziert. Diese Untersuchung bestätigt die prinzipielle Möglichkeit der elektronenmikroskopischen Ionenlokalisation durch simultane Gewebefixation und Ionenfällung nach der Methode von Komnick (1962). Die histochemische Lokalisation von Natrium- und Chlorionen im Schleim der apikalen Höhle der Chloridzellen von Stichlingen beweist die osmoregulatorisohe Punktion dieser Zellen und zeigt eine akkumulative Ionenadsorption durch die äußere Mucoidschicht an.

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On the histochemical localization of ions by electron microscopy, with special reference to the chloride reaction

Summary After dealing with the conditions, method and limitation of the histochemical localization of ions by means of electron microscopy several examples of chloride precipitation are given to explain some problems of interpretation and diffusion artifacts. The specifity of the histochemical method for chloride is demonstrated by control experiments on kidney, parietal cells and chloride cells. In chloride cells, the reaction product is identified by electron diffraction. This study principally confirms the possibility of the electron microscopical localization of ions by simultaneous tissue fixation and ion precipitation after the method of Komnick (1962).The histochemical localization of sodium and chloride in the mucus of the apical cavity of the stickleback chloride cells proves the osmoregulatory function of these cells and indicates an accumulative adsorption of ions by the extracellular mucous coat.

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Die Deutsche Forschungsgemeinschaft unterstützte die Untersuchungen durch eine Sachbeihilfe.     

Fine structure of chemosensitive cells (glomus caroticum) in tissue culture

Summary Cells of the carotid body of both embryonic and 1 to 2-d-old rabbits were cultured in monolayer in primary tissue culture. The cells grew in their original association of type I and type II cells. The fine structure of both cell types was similar to that in vivo. Even after one week in culture their morphological characteristics, such as the dense-cored vesicles, were preserved. The prolonged synthesis of catecholamines in culture and the formation of new granular vesicles is discussed.The results were reported in part at 17. Tagung für Elektronenmikroskopie held on September 21–26, 1975 in BerlinThe authors want to thank Prof. Dr. D.W. Lubbers for his advice and helpful suggestions     

人B细胞永生化研究进展

人源单克隆抗体具有免疫原性低、半衰期长等优势,成为了体内应用中不可或缺的生物制剂.人类抗体库为人源单克隆抗体的制备提供了丰富的来源,人B细胞永生化是获得人类抗体库的潜在有效方法,可应用于人源单克隆抗体的制备.由于各平台均有亟待解决的问题,基于人B细胞永生化的抗体制备尚局限在实验室研究阶段,且目前尚缺乏一篇系统综述以明确...     

The element distribution in ultrathin cryosections of cultivated fibroblast cells

A preparation method for measurements of the intracellular distribution of elements in tissue culture cells is described which is based on cryofixation, cryoultramicrotomy, cryotransfer and X-ray microanalysis in a scanning transmission electron microscope. Dry weight concentrations of phosphorus, sulfur, chlorine and potassium in the nucleus, the cytoplasm and the mitochondria of L929 fibroblast cells of the mouse are reported. The preparation and quantitation procedures are discussed with respect to present limitations and possible improvements of the method.     

Solid-phase extraction of phospholipids from hemoglobin solutions using Empore styrene–divinylbenzene disks

Styrene–divinylbenzene Empore disks were investigated for the extraction of phospholipids from red blood cells or aqueous solutions of hemoglobin as a means to reduce the time and solvent use required in sample preparation. Red blood cells are the source for hemoglobin used in the preparation of a hemoglobin-based oxygen carrier which is being developed to replace blood in transfusion therapy. Phospholipids are a major component of the membrane of red blood cells, and are toxic when administered directly into the vasculature. Sensitive analytical methods are required to detect phospholipids to ensure that concentrations in purified hemoglobin are well below toxic levels. This requires isolation from large volumes of purified hemoglobin solutions. The method described utilizes Empore disks to extract phospholipids from 30 ml of stroma free Hb preparations. Phosphatidylethanolamine, phosphatidylinositol, phosphatidylcholine and sphingomyelin were recovered with an average of 92% yield. The recovery of phosphatidylserine was 65%. The use of solvent and time required for sample preparation were reduced by an average of 80% relative to liquid–liquid extraction. The capacity of the 47-mm disk for the total of five phospholipids exceeds 0.3 mg. The method has been used for quantitation of phospholipids in red blood cells and stroma free hemoglobin solutions.     

Zur Funktionellen Morphologie der Speicheldrüsen Von Homopteren

Ohne ZusammenfassungIn Kurzform mitgeteilt auf der 53. Jahrestagung der Deutschen Zoologischen Gesellschaft in Münster i. W. (18.-23. 5. 59) und der 9. Tagung der Deutschen Gesellschaft für Elektronenmikroskopie in Freiburg i. Br. (18.-21. 10. 59).     

Besprechungen

Reviewed in this article:
Conners, I. L., An annotate index of plant diseases in Canada and fungi recorded on plants in Alaska.
Fahn, A., Plant Anatomy.
Mitteilungen aus der Biologischen Bundesanstalt fiir Land- und Forstwirtsdiaft BerMn-Dahlem, Heft 122 (1967).
Brandes, J., Elektronenmikroskopie von Pflanzenviren; Bibliographie von 1939–1965.
Müller, E. W., und H. J. Wasserburger, Insekten als Kulturpflanzenfeinde.     

Preparation of pre-cut corneas from fresh donated whole globes for Descemet’s stripping automated keratoplasty: 3-year results at the Central Eye Bank of Iran

To describe the technique and the results of the preparation of pre-cut corneas for Descemet’s stripping automated endothelial keratoplasty (DSAEK) during a 3-year period at the Central Eye Bank of Iran (CEBI). The method of preparation of pre-cut corneas from donated whole globes at the CEBI is described and the frequency and percentage of pre-cut corneas prepared for DSAEK, between April 2009 and March 2012, are specified. Moreover, post-operative reports are reviewed for any complaints about using pre-cut tissues for DSAEK. Out of the 1,518 donated whole globes appropriate for DSAEK, 1,478 (97.4 %) pre-cut corneas were successfully prepared. The method of preparation failed in 40 (2.6 %) cases. Based on the eye bank post-operative reports, thickness of pre-cut tissues for DSAEK was deemed unacceptable in only 6 (0.4 %) cases prior to surgery; five of these were too thick and one was too thin. Preparation of pre-cut corneas, for DSAEK from donated whole globes, in the CEBI is a safe and easy method, with very good preservation of endothelial cells after the preparation of the pre-cut corneas and reduced risks from corneal manipulation.     

Effective PCR detection of vegetative and dormant bacterial cells due to a unified method for preparation of template DNA encased within cell envelopes

The unified method of template preparation for PCR in the form of DNA covered by permeabilized cell envelopes was used for the cells of different physiological status (vegetative, dormant forms of different types, and nonviable micromummies). The procedure for the preparation of template DNA included one-stage (boiling in a buffer with chaotropic salts) or two-stage (boiling in a buffer with chaotropic salts followed by treatment with proteinase K) sample preparation. The proposed method proved effective for detection of not only vegetative cells but also of the bacillary spores and the cystlike dormant cells (CLC) of non-spore-forming bacteria. For example, the two-stage sample preparation of Bacillus cereus spores resulted in the PCR sensitivity increase up to the detection level of 3–30 spores per sample; the one-stage sample preparation was three orders of magnitude less efficient (10₄ spores per sample). An increase in the sensitivity of PCR detection (4–10-fold) owing to the use of the two-stage sample preparation was shown for bacillary, staphylococcal, and mycobacterial CLC. The possibility of PCR detection of staphylococcal micromummies with irreversibly lost viability, which were therefore undetectable by plating techniques, was also demonstrated. The application of the unified sample preparation method ensuring efficacious PCR detection of bacterial cells, irrespective of their physiological state, may be a promising approach to more complete detection of microbial diversity and the overall insemination of natural substrates.     

Mapping Goblet Cells in Mucous Membranes

A method is described for determining the number of goblet cells of the villi and crypts of Lieberkuhn in the small intestine with an accuracy far exceeding that which appears to be possible by counting on tissue sections. In groups of intact villi or crypts, previously isolated by microdissection, the goblet cells are stained, with as little staining as possible of the other tissue elements; thereafter the preparations are made transparent by embedding in a medium possessing a refractive index similar to that of the tissue. The staining is performed by the McManus-Hotchkiss periodic-leucofuchsin method (1948) with the modification that SchifPs reagent is diluted with 3 parts of water, the staining period cut down to 2V4-3 minutes, and the rinsing with bisulfite solution to 4-6 minutes. The embedding medium consists of colophonium and quinine hydrochloride in anise oil (Aurell, 1938). By this procedure, all the stained cells of the preparation may be visualized by manipulating the fine adjustment of the microscope. Counting of the goblet cells of the villi may be performed with great accuracy by projecting the picture of the preparation from the microscope on sectional paper and placing dots in the positions of the stained cells. The degree of magnification is determined by a corresponding projection of the scale of a micrometer disc.     

In situ chromosome preparation technique for simultaneous cytogenetic and immunocytochemical studies on cell cultures of solid tumors

Summary Immunophenotyping of cultured cancer cells requires intact antigenic structures; these are mostly destroyed by conventional chromosome preparation techniques. Thus, the simultaneous cytogenetic and immunocytochemical characterization of solid tumor cells appears unfeasible. Here, we describe a novel method that allows in situ chromosome preparation from monolayer cultures of solid tumor cells without affecting their immunological features. Using this technique, it is possible to achieve detailed cytogenetic data including chromosome banding together with the demonstration of cytoplasmic and nuclear antigens within the same tumor cell.     

Escherichia-erythrocyte diagnostic agents

The conditions of a simple and practicable method for the preparation of effective antigenic nonprotein diagnosticums on the basis of water-phenol extracts of 23 Escherichia species have been developed. The method consists in heating the mixture of erythrocytes and the antigen in a boiling water bath for 60 minutes. The diagnosticums thus obtained are 16-30 times more sensitive in the passive hemagglutination test and 4-6 times more sensitive in the passive hemagglutination inhibition test than diagnosticums prepared with the use of tannin, rivanol, as well as by the common method for the preparation of nonprotein antigens. The minimum concentration of Escherichia cells detected in the passive hemagglutination inhibition test is 0.8-1.2 million cells/ml.     

动物细胞膜的分离纯化及其在病毒受体研究中的应用

细胞膜是动物细胞与胞外环境之间的屏障。病毒只有与细胞膜上的病毒受体特异性结合 ,才能进入细胞 ,进而启动其增殖周期。因此 ,病毒受体是病毒学研究的重要组成部分。分离纯化病毒受体所在的细胞膜作为病毒受体研究的实验材料 ,已经在许多病毒的研究中得到应用 ,并取得了很好的效果。现就动物细胞膜的分离纯化及其在病毒受体研究中的应用作一综述。     

Discrepancies in ploidy determination due to specimen sampling errors

Two techniques are described to enhance the detection of low frequency aneuploid cells in automated cell analysis. One method concerns a cell preparation technique; the other is focused on specific cell selection at the measurement level. The cell preparation method has been designed to select and process the tumour areas in paraffin blocks and can be used for image as well as for flow cytometry. The technique uses incident fluorescence microscopy for visual inspection of the surface of the fluorescently stained tissue block to select the specific tumour parts. Using image cytometry, it is shown that in tissue sections with very small tumour foci and many normal cells, aneuploidy could only be detected after enrichment of the cell sample with the specifically selected areas. The cell selection at the measurement level is directed towards detection of low frequency aneuploid cells on microscope slides using the specific capacities of LEYTAS (Leyden Television Analysis System). With this system, cells of interest can be selected by means of minimum size and intensity thresholds. In addition to measurement of the total cell population, all cells above a minimum DNA value can thus be specifically selected and measured. The advantage of both enrichment techniques is the possibility to detect and measure aneuploid cell lines in cases where normal, diploid cells dominate the paraffin tissue.     

Experimental cloning of embryos through human-rabbit inter-species nuclear transfer

Therapeutic cloning,which is based on human somatic cell nuclear transfer,is one of our major research objectives.Though inter-species nuclear transfer has been introduced to construct human somatic cell cloned embryos,the effects of type,passage,and preparation method of donor cells on embryo development remain unclear.In our experiment,cloned embryos were reconstructed with different passage and preparation methods of ossocartilaginous cell,skin fibroblast,and cumulus cells.The cumulus cell embryos showed significantly higher development rates than the other two (P＜0.05).The development rate of embryos reconstructed with skin fibroblasts of different passage number and somatic cells of different chilling durations showed no significant difference.Also,fluorescence in situ hybridization (FISH)was conducted to detect nuclear derivation of the embryos.The result showed that the nuclei of the inter-species cloned embryo cells came from human.We conclude that (1)cloned embryos can be constructed through human-rabbit interspecies nuclear transfer;(2)different kinds of somatic cells result in different efficiency of nuclear transfer,while in vitro passage of the donor does not influence embryo development;(3)refrigeration is a convenient and efficient donor cell preparation method.Finally,it is feasible to detect DNA gcnotype through FISH.     

Experimental cloning of embryos through human-rabbit inter-species nuclear transfer

Therapeutic cloning, which is based on human somatic cell nuclear transfer, is one of our major research objectives. Though inter-species nuclear transfer has been introduced to construct human somatic cell cloned embryos, the effects of type, passage, and preparation method of donor cells on embryo development remain unclear. In our experiment, cloned embryos were reconstructed with different passage and preparation methods of ossocartilaginous cell, skin fibroblast, and cumulus cells. The cumulus cell embryos showed significantly higher development rates than the other two (P < 0.05). The development rate of embryos reconstructed with skin fibroblasts of different passage number and somatic cells of different chilling durations showed no significant difference. Also, fluorescence in situ hybridization (FISH) was conducted to detect nuclear derivation of the embryos. The result showed that the nuclei of the inter-species cloned embryo cells came from human. We conclude that (1) cloned embryos can be constructed through human-rabbit interspecies nuclear transfer; (2) different kinds of somatic cells result in different efficiency of nuclear transfer, while in vitro passage of the donor does not influence embryo development; (3) refrigeration is a convenient and efficient donor cell preparation method. Finally, it is feasible to detect DNA genotype through FISH. Translated from Zoological Research, 2005, 26(4): 416–421 [译自: 动物学研究]     

人-兔异种核移植构建克隆胚的实验研究

“治疗性克隆”是人类最关注的课题之一,而人体细胞核移植是治疗性克隆的基础和前提。异种核移植的方法虽已被引入人体细胞克隆胚的构建,但供体细胞的类型、培养代数及准备方法与其效率之间的关系尚有待探讨。本实验以不同培养代数和不同准备方法的人卵丘细胞、皮肤成纤维细胞和软骨细胞为供体构建了克隆胚,对其发育情况的比较表明,以卵丘细胞为供体时重构胚的体外发育率高于其余二者,差异显著（P〈0．05）;不同培养代数的成纤维细胞克隆胚和不同冷藏天数供体细胞克隆胚体外发育率无明显差异。此外,本实验还尝试用荧光原位杂交法检测所构建的异种克隆胚核遗传物质的来源,结果显示来自人体细胞。本研究表明,人一兔异种核移植构建克隆胚切实可行;体细胞的类型与核移植效率相关;供体细胞的体外培养传代对克隆胚的发育并无影响;而冷藏是一种简便有效的供体细胞准备方法;此外,用FISH方法对重构胚进行核遗传物质的鉴定切实可行。