Protocols in Current Issue
    Non-separate Mouse Sclerochoroid/RPE/Retina Staining and Whole Mount for the Integral Observation of Subretinal Layer
    Authors:  Sung-Jin Lee and Soo-Young Kim, date: 01/05/2021, view: 217, Q&A: 0

    The subretinal layer between retinal pigment epithelium (RPE) and photoreceptors is a region involved in inflammation and angiogenesis during the procession of diseases such as age-related macular degeneration. The current protocols of whole mounts (retina and RPE) are unable to address the intact view of the subretinal layer because the

    Assessing Self-interaction of Mammalian Nuclear Proteins by Co-immunoprecipitation
    Authors:  Claudia Cattoglio, Iryna Pustova, Xavier Darzacq, Robert Tjian and Anders S. Hansen, date: 02/20/2020, view: 2536, Q&A: 0
    [Abstract] Protein-protein interactions constitute the molecular foundations of virtually all biological processes. Co-immunoprecipitation (CoIP) experiments are probably the most widely used method to probe both heterotypic and homotypic protein-protein interactions. Recent advances in super-resolution microscopy have revealed that several nuclear proteins ...
    Immunohistochemical Identification of Muscle Fiber Types in Mice Tibialis Anterior Sections
    Authors:  Vanitha V. Rao and Abhishek Mohanty, date: 10/20/2019, view: 2491, Q&A: 0
    [Abstract] Mammalian skeletal muscle is a metabolically active tissue that is made up of different types of muscle fibers. These myofibers are made up of important contractile proteins that provide force during contraction of the muscle like actin and myosin. Murine myofibers have been classified into 4 types: Type I, Type IIa, Type IIb and Type IIX. Each ...
    A Highly Sensitive, Reproducible Assay for Determining 4-hydroxynonenal Protein Adducts in Biological Material
    Authors:  T. Blake Monroe and Ethan J. Anderson, date: 10/05/2019, view: 1312, Q&A: 0
    [Abstract] Oxidative stress is associated with numerous diseases, and markers of oxidative stress in biological material are becoming a mainstay of both experimental and clinical/epidemiological research. Lipid peroxidation is a major form of oxidative stress, but due to their rapid degradation and instability, lipid peroxides are notoriously difficult to ...
    Capillary Nano-immunoassay for Quantification of Proteins from CD138-purified Myeloma Cells
    Authors:  Irena Misiewicz-Krzeminska, Isabel Isidro and Norma C. Gutiérrez, date: 06/20/2019, view: 3308, Q&A: 0
    [Abstract] Protein analysis in bone marrow samples from patients with multiple myeloma (MM) has been limited by the low concentration of proteins obtained after CD138+ cell selection. A novel approach based on capillary nano-immunoassay could make it possible to quantify dozens of proteins from each CD138+ purified MM sample in an ...
    Measurement of CD74 N-terminal Fragment Accumulation in Cells Treated with SPPL2a Inhibitor
    Authors:  Rubén Martínez-Barricarte, Xiao-Fei Kong and Jean-Laurent Casanova, date: 06/05/2019, view: 2808, Q&A: 0
    [Abstract] The recent discovery of human signal peptide peptidase-like 2a (SPPL2a) deficiency in humans revealed the toxicity associated with the accumulation of one of its substrates, CD74 N-terminal fragment (CD74-NTF), for certain type of dendritic cells (cDC2). We developed a two-step protocol for monitoring the accumulation of this molecule in different ...
    Quantification of Cutaneous Ionocytes in Small Aquatic Organisms
    Authors:  Garfield T. Kwan, Shane H. Finnerty, Nicholas C. Wegner and Martin Tresguerres, date: 05/05/2019, view: 2623, Q&A: 0
    [Abstract] Aquatic organisms have specialized cells called ionocytes that regulate the ionic composition, osmolarity, and acid/base status of internal fluids. In small aquatic organisms such as fishes in their early life stages, ionocytes are typically found on the cutaneous surface and their abundance can change to help cope with various metabolic and ...
    Biofilm Assays on Fibrinogen-coated Silicone Catheters and 96-well Polystyrene Plates
    Authors:  Cristina Colomer-Winter, José A. Lemos and Ana L. Flores-Mireles, date: 03/20/2019, view: 3532, Q&A: 0
    [Abstract] Biofilm formation is a well-known bacterial strategy that protects cells from hostile environments. During infection, bacteria found in a biofilm community are less sensitive to antibiotics and to the immune response, often allowing them to colonize and persist in the host niche. Not surprisingly, biofilm formation on medical devices, such as ...
    Flow Cytometry Assay for Recycling of LFA-1 in T-lymphocytes
    Authors:  Katarzyna Potrzebowska, Janne Lehtonen, Malin Samuelsson and Lena Svensson, date: 12/05/2018, view: 2569, Q&A: 0
    [Abstract] To enable cells to move forward, cell surface integrins are internalized into an endosomal compartment and subsequently intracellularly transported to be re-exposed at a new site on the cell membrane. Leukocytes are the fastest migrating cell type in the human body, which express the leukocyte-specific integrin LFA-1. Here, we describe a flow ...
    Separation and Visualization of Low Abundant Ubiquitylated Forms
    Authors:  Ramona Schuster, Tânia Simões, Fabian den Brave and Mafalda Escobar-Henriques, date: 11/20/2018, view: 2542, Q&A: 0
    [Abstract] In this protocol we describe the separation and visualization of ubiquitylated forms of the yeast mitofusin Fzo1 by Western blot. To this aim, we express HA-tagged Fzo1 in Saccharomyces cerevisiae, break the cells to extract a membrane-enriched fraction, solubilize the membranes using detergent and then specifically immunoprecipitate the ...

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