Cell Biology

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    Protocols in Current Issue
    Analysis of Caenorhabditis elegans Sperm Number, Size, Activation, and Mitochondrial Content
    Authors:  Amy M. Hammerquist, Chia-An Yen and Sean P. Curran, date: 06/05/2021, view: 170, Q&A: 0
    [Abstract]

    Infertility is a widespread and often unexplained issue. Studying reproduction using C. elegans males offers insight into the influence of individual factors on male fertility in humans. We have created a collection of protocols to assess several aspects of C. elegans sperm quality, including number, size, rate of activation, and mitochondrial

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    Quantitation of Secretory Granule Size in Drosophila Larval Salivary Glands
    Authors:  Cheng-I J. Ma and Julie A. Brill, date: 06/05/2021, view: 161, Q&A: 0
    [Abstract]

    Maturation of secretory granules is a crucial process that ensures the bioactivity of cargo proteins undergoing regulated secretion. In Drosophila melanogaster, the larval salivary glands produce secretory granules that are up to four-fold larger in cross-sectional area after maturation. Therefore, we developed a live imaging microscopy approach

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    Visualization and Quantitation of Wg trafficking in the Drosophila Wing Imaginal Epithelium
    [Abstract]

    Secretory Wnt trafficking can be studied in the polarized epithelial monolayer of Drosophila wing imaginal discs (WID). In this tissue, Wg (Drosophila Wnt-I) is presented on the apical surface of its source cells before being internalized into the endosomal pathway. Long-range Wg secretion and spread depend on secondary secretion from endosomal

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    In vivo Fluorescence Imaging of Extracellular ATP in the Mouse Cerebral Cortex with a Hybrid-type Optical Sensor
    [Abstract]

    Adenosine 5’-triphosphate (ATP) works as an extracellular signaling molecule for cells in the brain, such as neurons and glia. Cellular communication via release of ATP is involved in a range of processes required for normal brain functions, and aberrant communication is associated with brain disorders. To investigate the mechanisms

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    Imaging and Fluorescence Quantification in Caenorhabditis elegans with Flow Vermimetry and Automated Microscopy
    Authors:  Elissa Tjahjono, Alexey V. Revtovich and Natalia V. Kirienko, date: 05/20/2021, view: 611, Q&A: 0
    [Abstract]

    Gene activation and cellular biomarkers are commonly monitored using fluorescent signals from transgenic reporters or dyes. These quantifiable markers are critical for biological research and serve as an incredibly powerful tool, even more so when combined with high-throughput screening. Caenorhabditis elegans is a particularly useful model in

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    A Workflow for High-pressure Freezing and Freeze Substitution of the Caenorhabditis elegans Embryo for Ultrastructural Analysis by Conventional and Volume Electron Microscopy
    Authors:  Mohammad M. Rahman, Irene Y. Chang, Orna Cohen-Fix and Kedar Narayan, date: 04/05/2021, view: 644, Q&A: 0
    [Abstract]

    The free-living nematode Caenorhabditis elegans is a popular model system for studying developmental biology. Here we describe a detailed protocol to high-pressure freeze the C. elegans embryo (either ex vivo after dissection, or within the intact worm) followed by quick freeze substitution. Processed samples are suitable for ultrastructural

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    Retention Using Selective Hooks (RUSH) Cargo Sorting Assay for Live-cell Vesicle Tracking in the Secretory Pathway Using HeLa Cells
    Authors:  Mehrshad Pakdel, Natalia Pacheco-Fernandez and Julia von Blume, date: 03/20/2021, view: 1075, Q&A: 0
    [Abstract]

    More than 30% of the total amount of proteins synthesized in mammalian cells follow the secretory pathway in order to mature and be properly sorted to their final destinations. Among several methodologies that describe live-cell monitoring of vesicles, the Retention Using Selective Hooks (RUSH) system is a powerful one that allows to visualize

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    Investigate Synaptic Vesicles Mobility in Neuronal Culture Axons by FRAP Imaging
    Authors:  Xiao Min Zhang, Fabrice P. Cordelieres and Etienne Herzog, date: 03/20/2021, view: 954, Q&A: 0
    [Abstract]

    Synaptic vesicles (SVs) are clustered in the presynaptic terminals and consistently trafficking along axons. Based on their release features, SVs are classified into different “pools”. Imaging of SVs that are traveling among multiple presynaptic terminals has helped define a new pool named “SV super-pool”. Here we describe

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    Retention Using Selective Hooks (RUSH) Cargo Sorting Assay for Protein Vesicle Tracking in HeLa Cells
    Authors:  Natalia Pacheco-Fernandez, Mehrshad Pakdel and Julia von Blume, date: 03/05/2021, view: 1548, Q&A: 0
    [Abstract]

    Monitoring vesicle trafficking is an excellent tool for the evaluation of protein dynamics in living cells. Such study is key for the understanding of protein sorting and secretion. Recent developments in microscopy, as well as new methodologies developed to study synchronized trafficking of proteins, allowed a better understanding of signaling,

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    Live Intravital Intestine with Blood Flow Visualization in Neonatal Mice Using Two-photon Laser Scanning Microscopy
    [Abstract]

    This protocol describes a novel technique to investigate the microcirculation dynamics underlying the pathology in the small intestine of neonatal mice using two-photon laser-scanning microscopy (TPLSM). Recent technological advances in multi-photon microscopy allow intravital analysis of different organs such as the liver, brain and intestine.

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