Topic
Lysis
About: Lysis is a research topic. Over the lifetime, 6072 publications have been published within this topic receiving 216978 citations.
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15 Sep 2004
TL;DR: Methods of Labeling Antibodies Detection of Antibody Binding Immunochemical Techniques: a Wide Range of Sensitive Assays for Detecting and Quantitating Antigen-Antibody Reactions Immunoprecipitation Immunohistochemistry Flow Cytometry.
Abstract: EXTRACTION OF PROTEIN Preparation of Buffers for Protein Extraction Use of Protease Inhibitors in Protein Extraction Use of Detergents in Protein Extraction Chemical Lysis for Protein Extraction Mechanical Lysis for Protein Extraction Preparation of Extract from Prokaryotes Extraction of Recombinant Protein from Bacteria Preparation of Extracts from Yeast Preparation of Extracts from Eukaryotes Preparation of Extracts from Plants Preparation of Membrane Extracts ESTIMATION OF PROTEIN Ultraviolet Absorption Methods Colorimetric Methods Fluorescent Methods ELECTROPHORETIC ANALYSES OF PROTEIN Driving Force of Electrophoresis Polyacrylamide Gel Electrophoresis (PAGE) Isoelectric Focusing (IEF) Two-Dimensional (2-D) Gel Electrophoresis Western Blotting Capillary Electrophoresis PURIFICATION OF PROTEIN General Consideration and Purification Strategy Non-Chromatographic Purification of Proteins Chromatographic Purification of Proteins ANTIBODIES: STRUCTURES, INTERACTIONS AND PRODUCTION Immune System and Antibody Response Structure of Antibodies Antigen-Antibody Interactions Production oof Antibodies Development of Monoclonal Antibodies Purification of Antibodies ANTIBODY LABELING, ANTIBODY DETECTION AND IMMUNOCHEMICAL TECHNIQUES Methods of Labeling Antibodies Detection of Antibody Binding Immunochemical Techniques: a Wide Range of Sensitive Assays for Detecting and Quantitating (Semi) Antigen-Antibody Reactions Immunoprecipitation Immunohistochemistry Flow Cytometry PURIFICATION OF GLYCOPROTEINS AND ANALYSES OF THEIR OLIGOSACCHARIDES Diagrams and Stereochemistry of Monosaccharides and Oligosaccharides Purification of Glycoproteins Release of Oligosaccharides From Glycoproteins Preparation of Glycopeptides by Proteolytic Cleavage Estimation of Carbohydrates Chromatographic Separation and Detection of Sugars Electrophoretic Separation Glycan Detection/Differentiation by Lectins Analyses of Carbohydrate Linkage
125 citations
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TL;DR: Strain PCP-1 was detected shortly after introduction into three soils at about the inoculum levels, as evidenced by both approaches, and results obtained by MPN-PCR with directly and indirectly extracted DNA were internally consistent.
Abstract: Since Mycobacterium chlorophenolicum strain PCP-1 is not detectable in soil by selective plating, a specific tracking method was based on the polymerase chain reaction (PCR) using soil DNA as a target A direct extraction protocol based on bead beating was adapted and used to obtain PCR-amplifiable DNA from five different soils In one soil, the disruption of cells of PCP-1, of Pseudomonas fluorescens R2f and of Paenibacillus azotofixans P3L5, as well as of the indigenous bacteria increased with increasing bead beating times After 45 min, lysis efficiency was about 90% or more in all cases Total DNA yields varied between soils, from 2 to 35 μg g–1 The purification steps needed to obtain amplifiable DNA were different per soil To detect target DNA specifically in bacterial cells, a new indirect extraction protocol was developed, which efficiently dislodged bacterial cells from the soil matrix, and produced amplifiable DNA with high yield To detect strain PCP-1 in soil, 16S ribosomal gene-based PCR combined with oligonucleotide hybridization was applied using a most-probable-number (MPN) set-up, whereas immunofluorescence was used for calibration Strain PCP-1 was detected shortly after introduction into three soils at about the inoculum levels, as evidenced by both approaches Both the direct and indirect DNA extraction methods yielded similar MPN estimates The dynamics of M chlorophenolicum PCP-1 was estimated in two soils over 14 days via MPN-PCR/oligonucleotide probing PCP-1 showed good survival in both soils, and results obtained by MPN-PCR with directly and indirectly extracted DNA were internally consistent Immunofluorescence cell enumerations supported the gross stability of PCP-1 in these two as well as in two additional soils
125 citations
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TL;DR: Ring-shaped membrane lesions generally similar to, but larger than, those previously described for complement lysis were observed, in agreement with recent measurements of larger functional pores for ADCC than complement.
Abstract: To test the hypothesis that complement-mediated cell lysis and cell-mediated cytotoxicity operate by analogous mechanisms, cell membranes from two antibody-dependent cytotoxicity systems were examined by electron microscopy after negative staining. Ring-shaped membrane lesions generally similar to, but larger than, those previously described for complement lysis were observed. These findings are in agreement with recent measurements of larger functional pores for ADCC than complement.
125 citations
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TL;DR: The results support the view that lysis of most target cells by cloned CTLs is due primarily to target cell membrane changes that are fundamentally equivalent to the formation of nonspecific ion channels.
Abstract: To investigate the destruction of target cells by murine CTLs, we examined intracellular Ca2+ concentrations ([Ca2+]i) and DNA fragmentation in target cells. Changes in [Ca2+]i were followed by flow cytometry by loading the cells with indo-1, a Ca2+-binding fluorescent dye, and determining the ration of fluorescence intensities at 405 nm (emission maximum for Ca2+-bound dye) over 480 nm (emission maximum for the free dye). Within minutes after interacting with the cytolytic granule fraction that had been isolated from CTLs, [Ca2+]i in target cells was strikingly increased. A pronounced increase in [Ca2+]i was also observed in target cells when they were specifically recognized by intact CTLs. Since ionomycin, a Ca2+ ionophore, caused a similar increase in [Ca2+]i and lysed cells (provided that extracellular Ca2+ was present), it appears that a sustained high level of [Ca2+]i is cytolytic. In contrast with other cells, CTLs, which have been shown to be refractory to granule-mediated lysis and to be poor targets for other CTLs, did not manifest an elevation in [Ca2+]i when they were similarly loaded with indo-1 and treated with isolated granules. The characteristic cleavage of target cell DNA into nucleosome-sized fragments was also induced by isolated granules as well as by valinomycin, a K+ ionophore, but not by ionomycin. The results support the view that lysis of most target cells by cloned CTLs is due primarily to target cell membrane changes that are fundamentally equivalent to the formation of nonspecific ion channels. The resulting large increase in [Ca2+]i is probably responsible for target cell lysis; and changes in intracellular ion concentrations also appear to be responsible for DNA fragmentation, probably by activating endogenous target cell endonucleases.
125 citations
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TL;DR: It is proposed that pulmonary inflammation during pneumococcal pneumonia arises in large part from the interaction of the bacterial cell wall with complement and noncomplement-mediated host defenses.
Abstract: Using a rabbit model of experimental pneumonitis, the components on the surface of the pneumococcus that incite pulmonary inflammation were identified. Rabbits were challenged intratracheally with live pneumococci, capsular polysaccharide, purified cell walls, or cell wall subcomponents. Leukocytosis and elevation of protein concentration was quantitated in bronchial lavage fluid during the first 24 h after challenge. Of the pneumococcal surface components tested, cell wall preparations had the highest specific activity in inducing inflammation; abnormalities in bronchial lavage fluid cytochemistry appeared rapidly and in a dose-dependent manner. Cell wall building blocks and the products of penicillin-induced hydrolysis of the cell wall were also highly inflammatory, indicating that inflammation can be generated by disruption of the cell wall during lysis of bacteria by beta-lactam antibiotics. Administration of inhibitors of arachidonic acid metabolism suggested that inhibition of the lipoxygenase pathw...
125 citations