ISSN:1059-910X    

DOI:10.1002/jemt.1070310102

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1995

数据来源: WILEY

摘要:

AbstractIn the present review, we report on recent results obtained by in situ transferaseimmunogold techniques as to the ultrastructural distribution of DNA and RNA within the cell nucleus. Special emphasis is placed on the various nucleolar components and the various enigmatic structures of the extranucleolar region: interchromatin granules, coiled bodies, and simple nuclear bodies. These data are discussed in the light of our current understanding of the functional organization of the cell nucleus. © 1995 Wiley‐Liss, I

ISSN:1059-910X    

DOI:10.1002/jemt.1070310103

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1995

数据来源: WILEY

摘要:

AbstractA significant amount of new information on structure‐function relationships in nuclei of adenovirus‐infected cells has accumulated during the last decade as a result of the combined use of several new cytochemical techniques. Localization of viral DNA on ultrathin sections of infected cells has been investigated at the ultrastructural level by using specific DNA staining and immunocytochemistry with monoclonal anti‐DNA antibodies. Both techniques, however, concomitantly visualize cellular and viral DNA. The specific stain for DNA reveals the configuration of the DNA molecules in the different nuclear substructures, whateer their synthetic activities. The immunodetection of DNA reveals that specific antibodies strongly bind to DNA of condensed host chromatin and to both encapsidated and nonencapsidated inactive viral genomes. However, the observation of an abnormally low level of labeling over the substructures in which synthetic activities of viral genomes are known to be intense demonstrates a serious limitation of this technique for the detection of active DNA. Postembedding in situ hybridization is the most useful method for identifying with certainty the structures containing defined nucleic acid sequences. By using a biotinylated viral DNA probe, in situ hybridization provides specific identification of structures containing either viral DNA or viral RNA molecules. In addition, with appropriate pretreatment of the sections, it is possible to reveal either all the viral DNA‐that is, both double‐ and single‐stranded DNA molecules (dsDNA, ssDNA)–or more specific species such as only ssDNA or only dsDNA molecules. The replicative and transcriptional activities of viral genomes are determined by high‐resolution autoradiography. Autoradiography after a short pulse incorporation of appropriate radioactive precursors by infected cells reveals the sites of cellular and viral DNA replication or trancription. A short pulse followed by chase periods of different durations reveals the progressive migration of the cellular and viral synthesized products. The in situ distribution of the viral 72 kDa DNA‐binding protein, a highly phosphorylated protein which protects the viral ssDNA, is revealed either by immunocytochemistry with specific antibodies or by the bismuth staining method which stains all highly phosphorylated proteins, including both cellular and viral proteins. The combined results of all these cytochemical procedures reveal the composition and functions of some of the structures induced by adenovirus infection. They demonstrate that viral genomes engaged in replication lead to the formation of replicative foci in which two compartments rapidly develop, one of which results from the aggregation of single strands of viral DNA and their accompanying 72 kDa protein. Conversely, ssDNA and 72 kDa protein are rare in the other compartment which is the main site of replication and transcription of viral genomes. The procedural aspects and the contributions of electron microscope cytochemistry to an understanding of the biology of Ad5 viruses can serve as a basic framework for the study of other biological systems. © 19

ISSN:1059-910X    

DOI:10.1002/jemt.1070310104

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1995

数据来源: WILEY

摘要:

AbstractImmunocytochemistry is a powerful tool for investigating protein secretion, extracellular matrix assembly, and cell‐matrix and matrix‐matrix/mineral relationships. When applied to the tissues of bones (bone and calcified cartilage) and teeth (dentin, cementum, and enamel), where calcium phosphate–containing extracellular matrices are the predominant structural component related to their weight‐bearing and masticatory roles, respectively, data from immunocytochemical studies have been prominent in advancing our understanding of mineralized tissue modeling and remodeling. The present review on the application of postembedding, colloidal‐gold immunocytochemistry to mineralized tissues focuses on the advantages of this approach and relates them to conceptual, theoretical, and experimental data currently available discussing matrix‐mineral interactions and extracellular matrix formation and turnover in these tissues. More specifically, data are summarized regarding the distribution and role of noncollagenous proteins in different mineralized tissues, particularly in the context of how they interface with mineral, and how this relationship might be affected by the various tissue‐processing steps and immunocytochemical strategies commonly implemented to examine the distribution and function of tissue proteins. Furthermore, a technical discussion is presented that outlines several different possibilities for epitope exposure in mineralized tissues during preparation of thin sections for transmission electron microscopy. Cell biological concepts of protein secretion by cells of the mineralized tissues, and subsequent extracellular matrix assembly and organization, are illustrated by examples of high‐resolution, colloidal‐gold immunolabeling for osteopontin, bone sialoprotein, and osteocalcin in the collagen‐based mineralized tissues and for enamel protein (amelogenin) in enamel. ©

ISSN:1059-910X    

DOI:10.1002/jemt.1070310105

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1995

数据来源: WILEY

摘要:

AbstractDuring the past few years, cyto‐ and immunocytochemical techniques have been developed and widely used for locating and identifying various molecules in plant cell compartments. The last decade has witnessed tremendous improvements in molecular cytology, thus allowing an accurate in situ detection of various components thought to play important biological functions in the plant metabolism. The use of immunocytochemistry to investigate resistance mechanisms of plants upon pathogen attack has provided key information on the defense strategy that plants elaborate during a host‐pathogen interaction. Of the various proteins induced in response to infection, chitinases and β‐1,3‐glucanases have been the focus of particular attention due to their believed antimicrobial activity through the hydrolysis of the main fungal wall components, chitin and β‐1,3‐glucans. Attention has also been paid to β‐fructosidase, the enzyme that hydrolyzes sucrose into glucose and fructoside. The marked accumulation of this enzyme upon pathogen infection has led to the consideration that infection may greatly infleunce the metabolic activity of colonized tissues by creating alterations of source‐sink relationships. Another facet of the plant's defense strategy that has been the focus of considerable interest is related to the accumulation of structural compounds, such as hydroxyproline‐rich glycoproteins and callose, to reinforce the wall architecture, thus decreasing vulnerability to microbial enzymes. A number of alternatives designed to improve plant protection towards pathogen invasion have been suggested. Among these, the production of transgenic plants expressing constitutively a foreign resistance gene and the pretreatment of plants with elicitors of defense reactions have been the subject of intensive studies at the molecular, biochemical, and cytological levels. Results of such studies clearly demonstrate the important contribution that cyto‐ and immunocytochemical approaches can make to our knowledge of how plants defend themselves and how plant disease resistance can be directly enhanced. These approaches will undoubtedly be active areas for future research in the development of biological control alternatives in which the mode of action of the product used is of key importance.

ISSN:1059-910X    

DOI:10.1002/jemt.1070310106

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1995

数据来源: WILEY

摘要:

AbstractPeroxisomes, since their discovery as microbodies, have been studied mostly independently by electron microscopists and biochemists. The fine structure has been studied by electron microscopy, and the compositional enzymes and proteins by protein biochemistry. Electron microscopic histochemistry has been used to try to clarify the relationship between the fine structure and its constituents. The immunogold technique, a combination of electron microscopy and protein biochemistry, for the first time resolved this problem due to the high sensitivity and resolution power of the staining and the high reliability of the technique. The present paper reviews the way in which the immunogold techniques, especially the protein A‐gold technique, revealed the localization of various enzymes or proteins in peroxisomes or peroxisomal subcompartments, and discusses why this technique should be employed in peroxisome research. © 1995 Wiley‐Liss,

ISSN:1059-910X    

DOI:10.1002/jemt.1070310107

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1995

数据来源: WILEY

ISSN:1059-910X    

DOI:10.1002/jemt.1070310108

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1995

数据来源: WILEY

ISSN:1059-910X    

DOI:10.1002/jemt.1070310101

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1995

数据来源: WILEY