Information value of the indices of the nonspecific resistance of the body

A number of nonspecific resistance characteristics in mice, such as the total number of peritoneal exudate cells, the percentage and absolute number of macrophages, their cytochemical activity in the spontaneous tetrazolium test and cytochemical capacity, have been studied by comparison with the resistance of the animals to tularemia infection induced by Francisella tularensis, Ga?ski?'s vaccinal strain 15. Of these characteristics, the cytochemical capacity of peritoneal exudate macrophages, i.e. the total cytochemical activity of macrophages contained in a unit of volume, has been the most informative as regards the level of nonspecific resistance to this infection. Other characteristics under study cannot serve as criteria for the evaluation of the nonspecific resistance of the body to F. tularensis.     

微观辨证学之研究：气血辨证与细胞化学含量的关系

在临床上为揭示肝脏病肝纤维化气虚血瘀和气滞血瘀病者,对细胞化学含量的变化,运用显微分光光度技术分析,结果发现气虚血瘀者单核细胞ＡＮＡＥ、ＡＣＰ,红细胞ＬＤＨ和嗜中性粒细胞ＡＤＰ明显降低,气滞血瘀者单核细胞ＡＮＡＥ、红细胞ＬＤＨ及嗜中性粒细胞ＡＤＰ较正常组偏低,但却较气虚血瘀组为高,单核细胞ＡＣＰ高于气虚血瘀组和正常对照组．据此,可将气虚、气滞血瘀及正常对照３组区别开来．     

Localization properties of fluorescence cytochemical enzyme procedures

Fluorescence enzyme cytochemical procedures will contribute significantly to biomedical problems where knowledge of the enzymic composition of individual cells is important. Compared with the number of absorbance enzyme cytochemical methods, relatively few fluorescence procedures have been reported. In this paper, the merits of the described methods are discussed. A distinction is made between methods with and without a capture reaction. Only a few methods satisfy the requirement of accurate localization of the final product and high signal to noise ratios. Thus, there still is a need for valid fluorescence cytochemical enzyme methods. It is concluded that the bottle neck for valid fluorescence cytochemical enzyme methods is the development of efficient fluorogenic capture reactions for the primary enzyme products.     

Histochemical features of the formation of the protective barrier of the stomach in various periods of ontogenesis of mammals

By means of a complex of histochemical methods it has been demonstrated that cytochemical differentiation of the tegmental epitheliocytes and secretory cells of the fundal glands takes place in different time. In Carnivores the tegmental epitheliocytes complete their cytochemical differentiation during the prenatal period, mucocytes--to the time of birth, and chief exocrinocytes--to the period of mixed feeding. In phytophagous animals formation of the protective barrier in the stomach occurs differently: in the mouse and hamster the cytochemical differentiation of the tegmental and glandular epitheliocytes is completed during the prenatal period, and in the rabbit and guinea pig--only by the 30th day after birth.     

Localization properties of fluorescence cytochemical enzyme procedures

Summary Fluorescence enzyme cytochemical procedures will contribute significantly to biomedical problems where knowledge of the enzymic composition of individual cells is important. Compared with the number of absorbance enzyme cytochemical methods, relatively few fluorescence procedures have been reported. In this paper, the merits of the described methods are discussed. A distinction is made between methods with and without a capture reaction. Only a few methods satisfy the requirement of accurate localization of the final product and high signal to noise ratios. Thus, there still is a need for valid fluorescence cytochemical enzyme methods. It is concluded that the bottle neck for valid fluorescence cytochemical enzyme methods is the development of efficient fluorogenic capture reactions for the primary enzyme products.In honour of Prof. P. van DuijnSupported (in part) by the Foundation for Medical Research (FUNGO), which is subsidized by the Netherlands Organization for the Advancement of Pure Research     

Cytochemical and biochemical microanalysis of carcinogenesis

Conclusions While the understanding of early cellular changes gleaned from conventional histopathology alone has been rather limited, modern cytochemical methods at the light and electron microscope level have revealed in a number of experimental models and in some human tumour types important results which indicate that a sequence of qualitatively different cell populations is followed during carcinogenesis Some of the cytochemical and electron microscope findings can be correlated with specific cytopathological phenomena which are readily detectable in routine H & E sections. Thus the evaluation of animal experiments concerned with the testing of chemical compounds for carcinogenicity has been improved. The use of simple cytochemical techniques may considerably increase the reliability of the results. With respect to the further elucidation of the mechanism of neoplastic cell transformation, cytochemistry has broadened research horizons. The identification of putative preneoplastic and early neoplastic cell populations by cytochemical methods allows for the first time the microdissection and subsequent detailed investigation of target cells of the carcinogen which are at a high risk of becoming cancer cells. The combination of cytochemical and biochemical microanalysis seems to be the most useful tool for clarifying a number of important problems of carcinogenesis at present.Dedicated to Professor Ekkehard Grundmann on the occasion of his sixtieth birthday.     

NSE/αNAE positivity in B-lineage acute lymphoblastic leukemia: revisiting a potential cytochemical diagnostic pitfall

Cytochemical staining for leukemia typing is declining in hematology laboratories, but the use of flow cytometry may not be possible in some settings. Aberrant cytochemical nonspecific esterase/α-naphthyl acetate esterase (NSE/αNAE) positive B-lymphoblasts can cause confusion with monoblasts, a potentially dangerous pitfall. This unusual cytochemical NSE/αNAE positivity had been associated with relatively poorer outcome of acute lymphoblastic leukemia (ALL) in the era prior to the advent of routine multicolor flow cytometric immunophenotyping. We reviewed morphological, cytochemical and flow-cytometric data from five cases of B-lineage ALL that showed NSE/αNAE positivity and were diagnosed definitively using multi-parametric flow cytometric immunophenotypic analysis. Diffuse or dot-like (localized) strong cytochemical NSE/αNAE activity was detected in all cases and all showed one or more features of high risk disease. The number of NSE/αNAE positive blasts in the marrow varied from 10 to 75%. The morphological differential diagnoses included T-lymphoid lineage ALL and acute monoblastic leukemia (AML-M5). Flow cytometric data revealed B-lineage antigens and the absence of monocytic or other myeloid markers resolved the diagnosis. These cases underscore the importance of immunophenotyping in all cases of suspected ALL regardless of the cytochemical findings. Although the numbers are small, the association with high risk disease observed in all five of our cases may corroborate the previously reported poor prognostic value of such aberrant cytochemical staining.     

Quantitative aspects of the cytochemical demonstration of glucose-6-phosphate dehydrogenase with tetranitro BT studied in a model system of polyacrylamide films

Summary The cytochemical determination of the activity of glucose-6-phosphate dehydrogenase (G6PDH) with tetranitro blue tetrazolium (TNBT) was studied with model films of polyacrylamide gel incorporating purified enzyme. This model system enabled a quantitative study to be made of different parameters involved with the cytochemical assay as it is applied to sections or smears. The enzyme activity of G6PDH incorporated in the model films was also assayed biochemically. Optimal conditions for retaining the maximum amount of enzymic activity are described. The behaviour of G6PDH towards enzyme inhibitors was found to be similar in model films and in solution. With TNBT, absorbance measurements at a single wavelength (535 nm) were used to estimate the enzyme activity quantitatively. When carried out under standardized conditions, both the cytochemical and biochemical assay showed a linear relation with the time of incubation and obeyed the Beer-Lambert law. The correlation between biochemical and cytochemical data was very high, which enabled cytochemical data to be converted into absolute units of enzyme activity. The data obtained in this way closely resembled the data of enzyme activity calculated from the absorbance of formazan produced inside polyacrylamide model films and afterwards extracted into a suitable solvent.     

植物酶的组织和细胞化学定位研究方法

酶是参与植物体内生化反应的特殊蛋白质。在保持活组织和细胞结构完整性的条件下,利用组织化学、细胞化学、免疫学和显微检测等技术研究酶的即位定位,是了解酶在组织、细胞和亚细胞中的分布、活性动态与定量及酶功能等的重要途径。对植物体中酶定位的组织化学和细胞化学方法的概念、原理与研究进展进行了综述,并根据国际酶化学分类编号顺序,分别介绍了25种酶的组织化学染色定位所用的反应介质和染色方法及46种酶的细胞化学定位方法的参考文献。     

Cytochemical study on the local tissue effect of nickel-containing intrauterine device in rabbits.

Authors introduced a nickel-containing device into the uterus of 20 rabbits, then performed the cytochemical study of the endometrium. During calcium cytochemical study in the endometrium intramitochondrial and partly intracytoplasmic localisation while with dimethylglyoxin method intracellular reaction precipitate were found. The nickel-containing device caused local nickel and intracellular or intramitochondrial calcium accumulation.     

In vitro cytochemical and cytophysiological study of in vivo BCG-activated mouse peritoneal macrophages

The morphological, cytochemical (acid phosphatase activity) and cytophysiological (phagocytosis) features of mouse peritoneal macrophages activated in vivo by BCG, were investigated in vitro. BCG induced an increase in cytochemical and phagocytic activities of the activated peritoneal macrophages.     

Granulocytic sarcoma with pleural involvement. Identification of neoplastic cells with cytochemistry

Malignant cells were detected in the pleural effusion of a patient with three separate primary malignancies. These cells were judged by conventional morphologic studies to be poorly differentiated cells, but cytochemical studies showed them to be granulocytic precursor cells. The use of cytochemical or immunochemical techniques may be most practical for the cytodiagnosis of malignant cells in serous effusions.     

Aminoglycoside binding sites in Escherichia coli as revealed by neomycin-gold labeling

A cytochemical technique for demonstration of neomycin binding sites by electron microscopy was developed and applied to Escherichia coli. Neomycin was conjugated chemically with bovine serum albumin (BSA). Colloidal gold was coated with the conjugated neomycin-BSA. The neomycin-BSA-gold was applied to thin sections of Epon-embedded E. coli and examined. Gold particles were observed on the outer membrane and the cytoplasmic membrane of E. coli. It was probably the ribosomes that were being labeled in the cytoplasm. Different cytochemical controls, including a number of inhibition tests and the use of BSA-gold, proved the specificity of this cytochemical technique and provided the biochemical significance of the observations.     

A new dynamic model system for the study of capture reactions for diffusable compounds in cytochemistry. II. Effect of the composition of the incubation medium on the trapping of phosphate ions in acid phosphatase cytochemistry

Synopsis A model system developed for the study of the dynamics of capture reactions for diffusable compounds in cytochemistry served as a basis for the experiments reported in the present paper. The model was used to study the effect of the composition of the cytochemical medium on the trapping of phosphate ions by lead (II) ions in acid phosphatase cytochemistry. In this system a phosphate-containing solution and a lead-containing solution (cytochemical medium) are pumped along opposite sides of a polyacrylamide film. The phosphate concentration at which measurable precipitation starts in the film (critical phosphate concentration) was taken as a measure of the trapping efficiency of the cytochemical medium. The addition of -glycerophosphate and cytidine-5-monophosphate to a buffered lead-containing solution resulted in a higher critical phosphate concentration. Both substrates had an effect on the crystal form of lead phosphate. The addition of chloride ions and acetone, as well as decreasing the molarity of the acetate buffer of the cytochemical medium, were found to lower the critical phosphate concentration, whereas the addition of fluoride ions, glucose, and sucrose had no effect. From the effect of variations in the composition of the cytochemical medium on the trapping efficiency and the turnover number of acid phosphatase in the medium, it was possible to predict which cytochemical medium would be the most suitable for the demonstration of acid phosphatase activity in guinea-pig peritoneal exudate cells. The results were in accordance with the localization of acid phosphatase activity: the higher the trapping efficiency and the turnover number, the higher the amount of precipitate and the number of positive enzymatic sites. In this way an improved cytochemical medium for acid phosphatase was developed.     

Cytochemical characterization of leucocytes from the seawater teleost,gilthead seabream (Sparus aurata L.)

The cytochemical characterization of head-kidney and peripheral blood leucocytes of gilthead seabream (Sparus aurata L.) was studied by light and electron microscopy. Neutrophilic granulocytes show some cytoplasmic granules, which are positive for alkaline phosphatase and peroxidase but acid phosphatase negative. The scarce granules found in the cytoplasm of the circulating neutrophils and their cytochemical features seem to be indicative of an immature stage. Acidophils are also alkaline phosphatase and peroxidase positive at pH 11.0. They are strongly positive for acid phosphatase and acid phosphatase activity may thus be considered a cytochemical marker to characterize and differentiate neutrophilic from acidophilic granulocytes in this fish species. Three granule populations are characterized in the cytoplasm of the gilthead seabream acidophils: the first is positive only for peroxidase and the second contains a dense core with acid and alkaline phosphatase activities, surrounded by a thin peroxidase positive electron-dense halo. The third granule type contains an eccentric core, which is strongly positive for acid and alkaline phosphatase and peroxidase. As regards their cytochemical features, the first and second granule types seem to correspond respectively to the azurophilic and specific granules found in acidophils of mammals and could be involved in phagocytic processes, thus playing an important microbicidal role in this species. The monocytes, monocyte-macrophages and macrophages show different cytochemical features. The first have scarce acid phosphatase-positive lysosomes, while blood monocyte-macrophages and macrophages are positive for acid and alkaline phosphatases and for peroxidase; the monocyte-macrophages show scarce lysosomes.     

Model film studies in enzyme histochemistry with special reference to glucose-6-phosphate dehydrogenase

Summary This paper deals with the progress made over the last few years in our understanding of enzyme cytochemical staining methods as studied using a fundamental approach with the aid of a model system of thin gel films. Although model films with a matrix of polyacrylamide have been mostly used, the properties and possible applications of other matrices are also reviewed. The chemical aspects of the entrapment of enzyme molecules into a matrix are summarized. Special attention has been paid in model film studies to the principles of the trapping reaction of a diffusable precursor resulting from the enzymatic conversion of a substrate. They are considered here as they concern the cytochemical demonstration of acid phosphatase activity with a lead salt. The effect of fixatives on different enzyme activities, the diffusion rate of substrates and chromogenic compounds to the enzyme site, and enzyme kinetics under cytochemical conditions are also discussed, since they are factors which influence the final results of the staining procedures. The advantage of model film studies in enabling the direct correlation of cytochemical and biochemical results is outlined with special reference to the cytochemical determination of glucose-6-phosphate dehydrogenase with Tetra Nitro BT. A method for determining enzyme activities in the soluble fraction of isolated cells after incorporation in model films is described for the first time. This method has proved to be highly appropriate for microscopical observations of glucose-6-phosphate dehydrogenase activity in single cells, because it results in a good morphology and no formazan precipitaties outside the cells. On the other hand, this type of model film forms a bridge between fundamental model film studies using purified enzyme and quantitative enzyme cytochemistry performedin situ.     

A new method for the enzyme cytochemical staining of individual cells with the use of a polyacrylamide carrier

A new method for enzyme cytochemical studies on individual cells is developed. Cells are incorporated in the matrix of a thin film of transparent polyacrylamide prior to incubation in a cytochemical medium. Five different kinds of individual cells, i.e. isolated rat hepatocytes, isolated mouse oocytes, cultivated human fibroblasts, rat thymocytes and human blood cells are used for testing the applicability of this method for the cytochemical demonstration of glucose-6-phosphate dehydrogenase with tetranitro BT. The incorporation technique solves at least some of the problems occurring with enzyme cytochemistry on single cells. The morphology of the cell is very well preserved, the formazan precipitation due to enzyme activity occurs entirely within the cell cytoplasm, the nothing dehydrogenase activity can be kept very low and the loss of cells is completely prevented with all cell types used.     

DIFFERENTIATION OF ENDOPLASMIC RETICULUM IN HEPATOCYTES : II. Glucose-6-Phosphatase in Rough Microsomes

Electron microscope cytochemical localization of glucose-6-phosphatase in the developing hepatocytes of fetal and newborn rats indicates that the enzyme appears simultaneously in all the rough endoplasmic reticulum of a cell, although asynchronously within the hepatocyte population as a whole. To confirm that the pattern of cytochemical deposits reflects the actual distribution of enzyme sites, a method to subfractionate rough endoplasmic reticulum was developed. The procedure is based on the retention of the cytochemical reaction product (precipitated lead phosphate) within freshly prepared rough microsomes reacted in vitro with glucose-6-phosphate and lead ions. Lead phosphate increases the density of the microsomes which have glucose-6-phosphatase activity and thereby makes possible their separation from microsomes lacking the enzyme; separation is obtained by isopycnic centrifugation on a two-step density gradient. The procedure was applied to rough microsomes isolated from rats at several stages during hepatocyte differentiation and the results obtained agree with those given by cytochemical studies in situ. Before birth, when only some of the cells react positively for glucose-6-phosphatase, only a commensurate proportion of the rough microsome fraction can be rendered dense by the enzyme reaction. At the time of birth and in the adult, when all cells react positively, practically all microsomes acquire deposit and become dense after reaction. Thus, the results of the microsome subfractionation confirm the cytochemical findings; the enzyme is evenly distributed throughout all the endoplasmic reticulum of a cell and there is no regional differentiation within the rough endoplasmic reticulum with respect to glucose-6-phosphatase. These findings suggest that new components are inserted molecule-by-molecule into a pre-existing structural framework. The membranes are thus mosaics of old and new molecules and do not contain large regions of entirely "new" membrane in which all of the components are newly synthesized or newly assembled.     

Cytochemical localization of acid phosphatase in striated muscle

Normal skeletal and cardiac striated muscle from adult rats was incubated for the cytochemical detection of acid phosphatase activity with cerium as the capture metal. Results from these experiments show that normal striated muscle has a greater number of acid phosphatase-positive structures, which are presumed to be lysosomes, than has been indicated by several previous cytochemical studies.