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    Differentiation of Myeloid-derived Suppressor Cells from Murine Bone Marrow and Their Co-culture with Splenic Dendritic Cells
    Authors:  Giada Mondanelli and Claudia Volpi, date: 09/20/2017, view: 30, Q&A: 0
    Myeloid-derived suppressor cells (MDSCs) possess the ability to suppress the immune response, and to amplify the regulatory properties of other immune cells, i.e., dendritic cells. Here we describe a protocol in which MDSCs were ...
    Phagocytosis Assay of Necroptotic Cells by Cardiac Myofibroblasts
    In myocardial infarction (MI), a plenty of cardiomyocytes undergo necrosis and necroptosis due to the lack of oxygen and nutrients. The dead cardiomyocytes are promptly engulfed by phagocytes. When the dead cells are not engulfed, the noxious ...
    An Assay to Determine Phagocytosis of Apoptotic Cells by Cardiac Macrophages and Cardiac Myofibroblasts
    In myocardial infarction (MI), a number of cardiomyocytes undergo apoptosis. These apoptotic cardiomyocytes are promptly engulfed by phagocytes. If the dead cells are not engulfed, their noxious contents are released outside, resulting in induction ...
    Freeze-fracture-etching Electron Microscopy for Facile Analysis of Yeast Ultrastructure
    Authors:  Takuma Tsuji and Toyoshi Fujimoto, date: 09/20/2017, view: 49, Q&A: 0
    We describe a streamlined method that enables the quick observation of yeast ultrastructure by electron microscopy (EM). Yeast cells are high-pressure frozen, freeze-fractured to cut across the cytoplasm, and freeze-etched to sublimate ice in the ...
    Preparation of Primary Cultures of Embryonic Rat Hippocampal and Cerebrocortical Neurons
    This protocol aims at standardizing the procedure to obtain primary cultures of hippocampal and cerebrocortical neurons for in vitro experiments. Cultures should be prepared from cells isolated during embryonic development when neuronal ...
    Uptake Assays to Monitor Anthracyclines Entry into Mammalian Cells
    Authors:  Nicolas Brosseau, Emil Andreev and Dindial Ramotar, date: 09/20/2017, view: 29, Q&A: 0
    Anthracyclines, such as doxorubicin and daunorubicin, are DNA damaging agents that autofluoresce and can be readily detected in cells. Herein, we developed suitable assays to quantify and localize daunorubicin in mammalian cells. These assays can be ...
    Differentiation of Myeloid-derived Suppressor Cells from Murine Bone Marrow and Their Co-culture with Splenic Dendritic Cells
    Authors:  Giada Mondanelli and Claudia Volpi, date: 09/20/2017, view: 30, Q&A: 0
    [Abstract] Myeloid-derived suppressor cells (MDSCs) possess the ability to suppress the immune response, and to amplify the regulatory properties of other immune cells, i.e., dendritic cells. Here we describe a protocol in which MDSCs were differentiated from murine bone marrow cells, and CD11c+ dendritic cells were purified from murine ...
    Phagocytosis Assay of Necroptotic Cells by Cardiac Myofibroblasts
    [Abstract] In myocardial infarction (MI), a plenty of cardiomyocytes undergo necrosis and necroptosis due to the lack of oxygen and nutrients. The dead cardiomyocytes are promptly engulfed by phagocytes. When the dead cells are not engulfed, the noxious contents of the cells are released outside, and thus, induce inflammation, and obstruct the function of ...
    An Assay to Determine Phagocytosis of Apoptotic Cells by Cardiac Macrophages and Cardiac Myofibroblasts
    [Abstract] In myocardial infarction (MI), a number of cardiomyocytes undergo apoptosis. These apoptotic cardiomyocytes are promptly engulfed by phagocytes. If the dead cells are not engulfed, their noxious contents are released outside, resulting in induction of inflammation. Therefore, the removal of these dead cells is necessary. However, the contribution ...
    Freeze-fracture-etching Electron Microscopy for Facile Analysis of Yeast Ultrastructure
    Authors:  Takuma Tsuji and Toyoshi Fujimoto, date: 09/20/2017, view: 49, Q&A: 0
    [Abstract] We describe a streamlined method that enables the quick observation of yeast ultrastructure by electron microscopy (EM). Yeast cells are high-pressure frozen, freeze-fractured to cut across the cytoplasm, and freeze-etched to sublimate ice in the cytosol and the organelle lumen. The cellular structures delineated by these procedures are coated by ...
    Preparation of Primary Cultures of Embryonic Rat Hippocampal and Cerebrocortical Neurons
    [Abstract] This protocol aims at standardizing the procedure to obtain primary cultures of hippocampal and cerebrocortical neurons for in vitro experiments. Cultures should be prepared from cells isolated during embryonic development when neuronal precursor cells are not yet fully differentiated. This helps increasing the quality and quantity of ...
    Uptake Assays to Monitor Anthracyclines Entry into Mammalian Cells
    Authors:  Nicolas Brosseau, Emil Andreev and Dindial Ramotar, date: 09/20/2017, view: 29, Q&A: 0
    [Abstract] Anthracyclines, such as doxorubicin and daunorubicin, are DNA damaging agents that autofluoresce and can be readily detected in cells. Herein, we developed suitable assays to quantify and localize daunorubicin in mammalian cells. These assays can be exploited to identify components that are involved in the uptake of anthracyclines.
    Peroxisome Motility Measurement and Quantification Assay
    Authors:  Jeremy Metz, Inês G. Castro and Michael Schrader, date: 09/05/2017, view: 185, Q&A: 0
    [Abstract] Organelle movement, distribution and interaction contribute to the organisation of the eukaryotic cell. Peroxisomes are multifunctional organelles which contribute to cellular lipid metabolism and ROS homeostasis. They distribute uniformly in mammalian cells and move along microtubules via kinesin and dynein motors. Their metabolic cooperation ...
    Protocol for Establishing a Multiplex Image-based Autophagy RNAi Screen in Cell Cultures
    Authors:  Jennifer Jung and Christian Behrends, date: 09/05/2017, view: 233, Q&A: 0
    [Abstract] Autophagy is a recycling pathway, in which intracellular cargoes including protein aggregates and bacteria are engulfed by autophagosomes and subsequently degraded after fusion with lysosomes. Dysregulation of this process is involved in several human diseases such as cancer or neurodegeneration. Hence, advancing our understanding of how autophagy ...
    Isolation and Separation of Epithelial CD34+ Cancer Stem Cells from Tgfbr2-deficient Squamous Cell Carcinoma
    Authors:  Heather A. McCauley and Géraldine Guasch, date: 09/05/2017, view: 256, Q&A: 0
    [Abstract] Most epithelial tumors have been shown to contain cancer stem cells that are potentially the driving force in tumor progression and metastasis (Kreso and Dick, 2014; Nassar and Blanpain, 2016). To study these cells in depth, cell isolation strategies relying on cell surface markers or fluorescent reporters are essential, and the isolation ...
    In vivo Priming of T Cells with in vitro Pulsed Dendritic Cells: Popliteal Lymph Node Assay
    Authors:  Songjie Cai, Masayuki Fujino, Lina Lu and Xiao-Kang Li, date: 09/05/2017, view: 182, Q&A: 0
    [Abstract] One-way mixed lymphocyte reaction (MLR) is a classic tool to measure how T cells react to external stimuli. However, MLR is an in vitro reaction system, which shows different response intensity compared with in vivo trails sometimes due to the lack of cytokines, tissue matrix and other immune response associated factors. The ...