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Large-Scale Mitotic Cell Synchronization

Understanding cell growth and cell division involves the study of regulatory events that occur in a cell cycle phase-dependent manner. Studies analyzing cell cycle regulatory mechanisms and cell cycle progression invariably require synchronization of cell populations at spec ...

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Synchronization of Mammalian Cells and Nuclei by Centrifugal Elutriation

Synchronized populations of large numbers of cells can be obtained by centrifugal elutriation on the basis of sedimentation properties of small round particles, with minimal perturbation of cellular functions. The physical characteristics of cell size and sedimentation veloc ...

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Overview of Cell Synchronization

Widespread interest in cell synchronization is maintained by the studies of control mechanisms involved in cell cycle regulation. During the synchronization distinct subpopulations of cells are obtained representing different stages of the cell cycle. These subpopulations ...

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Molecular Network Dynamics of Cell Cycle Control: Transitions to Start and Finish

The cell cycle is controlled by complex regulatory network to ensure that the phases of the cell cycle happen in the right order and transitions between phases happen only if the earlier phase is properly finished. This regulatory network receives signals from the environment, monitors the st ...

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Flow Cytometry Developments and Perspectives in Clinical Studies: Examples in ICU Patients

Septic syndromes represent a major, although largely under-recognized, healthcare problem worldwide accounting for thousands of deaths every year. Although flow cytometry (FCM) remains a relatively confidential diagnostic tool, it is useful at every step of intensive care unit ( ...

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Ex Vivo Expansion of Haematopoietic Stem Cells to Improve Engraftment in Stem Cell Transplantation

The efficient use of haematopoietic stem cells (HSC) for transplantation is often limited by the relatively low numbers of HSC collected. The ex vivo expansion of HSC for clinical use is a potentially valuable and safe approach to increase HSC numbers thereby increasing engraftment and redu ...

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Cell Cycle Synchronization for the Purpose of Somatic Cell Nuclear Transfer (SCNT)

Somatic cell nuclear transfer (SCNT) is a technically and biologically challenging procedure during which a differentiated committed nucleus undergoes rapid reprogramming into the totipotent state in a few hours. SCNT can be utilized to generate patient- and disease-specific em ...

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Synchronization of Medicago sativa Cell Suspension Culture

Deepening our knowledge on the regulation of the plant cell division cycle depends on techniques that allow for the enrichment of cell populations in defined cell cycle phases. Synchronization of cell division can be achieved using different plant tissues; however, well-established c ...

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Synchronization of In Vitro Maturation in Porcine Oocytes

When removed from the follicles, during the 44h process of in vitro maturation (IVM) fully grown porcine oocytes resume meiosis spontaneously from the late diplotene stage of the first meiotic prophase and proceed to the metaphase-II (MII) stage at which they remain arrested until fertiliz ...

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Synchronization of Pathogenic Protozoans

Protozoans are single-cell eukaryotes and many of the best studied protozoans are parasitic to humans (e.g., Plasmodium falciparum causing malaria and Trypanosoma brucei causing sleeping sickness). These organisms are distantly related to humans but with retained eukaryotic ty ...

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Synchronization of Yeast

The budding yeast Saccharomyces cerevisiae and fission yeast Schizosaccharomyces pombe are amongst the simplest and most powerful model systems for studying the genetics of cell cycle control. Because yeast grows very rapidly in simple and economical media, large numbers of cells can ...

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Synchronization of Bacillus subtilis Cells by Spore Germination and Outgrowth

This protocol defines conditions under which the germination of spores can be used to synchronize Bacillus subtilis cells, utilizing the time-ordered sequence of events taking place during the transition from spore to vegetative cells. The transition stages involve: phase change, s ...

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Synchronization of HeLa Cells

HeLa is one of the oldest and most commonly used cell lines in biomedical research. Owing to the ease of which they can be effectively synchronized by various methods, HeLa cells have been used extensively for studies of the cell cycle. Here we describe several protocols for synchronization of HeLa c ...

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Cytofluorometric Purification of Diploid and Tetraploid Cancer Cells

During malignant transformation, cells can increase their ploidy and hence become polyploid (mostly tetraploid). Frequently, however, tetraploid cells undergo asymmetric divisions, in turn entailing a reduction in ploidy and the acquisition of a pseudo-diploid, aneuploid st ...

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Semipermeabilized Cells to Study Procollagen Assembly

This chapter will describe the preparation and use of a semipermeabilized (SP) cell system that reconstitutes the initial stages in the assembly and modification of proteins entering the secretory pathway (1). The procedure involves treating cells grown in culture with the detergent d ...

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Atomic Force Microspopy Measurements of Intermolecular Binding Strength

Intermolecular binding forces are intrinsic property of cohesive structures and should be used as the main quantitative criteria for assessing and defining their functional contribution to the maintenance of the anatomical integrity of the adult and embryonal multicellular or ...

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ECM Macromolecules: Rotary Shadowing and Scanning Transmission Electron Microscopy

The examination of ECM macromolecules by conventional transmission electron microscopy (TEM) or scanning transmission EM (STEM) can provide important information on macromolecular organization and interactions.

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Quantitative Determination of Collagen Crosslinks

The primary functional role of collagen is as a supporting tissue and it is now well established that the aggregated forms of the collagen monomers are stabilized to provide mechanical strength by a series of intermolecular crosslinks. These links are formed by oxidative deamination of the ε- ...

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Analysis of Laminin Structure and Function with Recombinant Glycoprotein Expressed in Insect Cells

Recent developments in the application of eukaryotic recombinant protein techniques have provided new tools with which to dissect and map functional activities in basement membrane glycoproteins. This has been particularly valuable in the case of laminins where the relationsh ...

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Eukaryotic Expression and Purification of Recombinant Extracellular Matrix Proteins Carrying the Strep II Tag

For recombinant expression of extracellular matrix (ECM) proteins or their individual domains, the use of transformed mammalian cells offers two major advantages. First, eukaryotic expression can be expected under optimum conditions to produce a large proportion of correctly fo ...

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