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    Protocols in Current Issue
    Identification and Quantitation of Leukocyte Populations in Human Kidney Tissue by Multi-parameter Flow Cytometry
    Inflammatory immune cells play direct pathological roles in cases of acute kidney injury (AKI) and chronic kidney disease (CKD). However, the identification and characterization of distinct populations of leukocytes in human kidney biopsies have ...
    Quantitative ChIP-seq by Adding Spike-in from Another Species
    Authors:  Kongyan Niu, Rui Liu and Nan Liu, date: 08/20/2018, view: 49, Q&A: 0
    Chromatin immunoprecipitation followed by sequencing (ChIP-seq) is a routine procedure in the lab; however, epigenome-wide quantitative comparison among independent ChIP-seq experiments remains a challenge. Here, we contribute an experimental ...
    Studying the Role of Microglia in Neurodegeneration and Axonal Regeneration in the Murine Visual System
    Authors:  Alexander M Hilla and Dietmar Fischer, date: 08/20/2018, view: 43, Q&A: 0
    Microglia reside in the central nervous system (CNS) and are involved in the maintenance of the physiologic state. They constantly survey their environment for pathologic alterations associated with injury or diseases. For decades, researchers have ...
    Trace Fear Conditioning: Procedure for Assessing Complex Hippocampal Function in Mice
    The trace fear conditioning protocol is designed to measure hippocampal function in mice. The protocol includes a neutral conditioned stimulus (tone) and an aversive unconditioned stimulus (shock), separated in time by a trace interval. The trace ...
    Image-Based Analysis of Mitochondrial Area and Counting from Adult Mouse Dopaminergic Neurites
    Authors:  Nadee Nissanka, Carlos T Moraes and Mlena Pinto, date: 08/20/2018, view: 26, Q&A: 0
    Mitochondria form dynamic cytoplasmic networks which undergo morphological changes in order to adapt to cellular stresses and signals. These changes can include alterations in size and number within a given cell. Analysis of the whole network can be ...
    Generation of Human Mesenchymal Stem Cell 3D Spheroids Using Low-binding Plates
    The 3D culture of human mesenchymal stem cells (hMSCs) represents a more physiological environment than classical 2D culture and has been used to enhance the MSC secretome or extend cell survival after transplantation. Here we describe a simple and ...
    Deoxycholate Fractionation of Fibronectin (FN) and Biotinylation Assay to Measure Recycled FN Fibrils in Epithelial Cells
    Authors:  Archana Varadaraj, Carina Magdaleno and Karthikeyan Mythreye, date: 08/20/2018, view: 28, Q&A: 0
    Fibronectin (FN) is an extracellular matrix protein that is secreted by many cell types and binds predominantly to the cell surface receptor Integrin α5β1. Integrin α5β1 binding initiates the step-wise assembly of FN into fibrils, a process called ...
    Identification and Quantitation of Leukocyte Populations in Human Kidney Tissue by Multi-parameter Flow Cytometry
    [Abstract] Inflammatory immune cells play direct pathological roles in cases of acute kidney injury (AKI) and chronic kidney disease (CKD). However, the identification and characterization of distinct populations of leukocytes in human kidney biopsies have been confounded by the limitations of immunohistochemical (IHC)-based techniques used to detect them. ...
    Quantitative ChIP-seq by Adding Spike-in from Another Species
    Authors:  Kongyan Niu, Rui Liu and Nan Liu, date: 08/20/2018, view: 49, Q&A: 0
    [Abstract] Chromatin immunoprecipitation followed by sequencing (ChIP-seq) is a routine procedure in the lab; however, epigenome-wide quantitative comparison among independent ChIP-seq experiments remains a challenge. Here, we contribute an experimental protocol combined with a computational workflow allowing quantitative and comparative assessment of ...
    Studying the Role of Microglia in Neurodegeneration and Axonal Regeneration in the Murine Visual System
    Authors:  Alexander M Hilla and Dietmar Fischer, date: 08/20/2018, view: 43, Q&A: 0
    [Abstract] Microglia reside in the central nervous system (CNS) and are involved in the maintenance of the physiologic state. They constantly survey their environment for pathologic alterations associated with injury or diseases. For decades, researchers have investigated the role of microglia under different pathologic conditions, using approaches aiming to ...
    Trace Fear Conditioning: Procedure for Assessing Complex Hippocampal Function in Mice
    [Abstract] The trace fear conditioning protocol is designed to measure hippocampal function in mice. The protocol includes a neutral conditioned stimulus (tone) and an aversive unconditioned stimulus (shock), separated in time by a trace interval. The trace interval between the tone and the shock critically involves the hippocampus and could be used to ...
    Image-Based Analysis of Mitochondrial Area and Counting from Adult Mouse Dopaminergic Neurites
    Authors:  Nadee Nissanka, Carlos T Moraes and Mlena Pinto, date: 08/20/2018, view: 26, Q&A: 0
    [Abstract] Mitochondria form dynamic cytoplasmic networks which undergo morphological changes in order to adapt to cellular stresses and signals. These changes can include alterations in size and number within a given cell. Analysis of the whole network can be a useful metric to assess overall mitochondrial health, particularly in neurons, which are highly ...
    Generation of Human Mesenchymal Stem Cell 3D Spheroids Using Low-binding Plates
    [Abstract] The 3D culture of human mesenchymal stem cells (hMSCs) represents a more physiological environment than classical 2D culture and has been used to enhance the MSC secretome or extend cell survival after transplantation. Here we describe a simple and affordable method to generate 3D spheroids of hMSCs by seeding them at high density in a low-binding ...
    Deoxycholate Fractionation of Fibronectin (FN) and Biotinylation Assay to Measure Recycled FN Fibrils in Epithelial Cells
    Authors:  Archana Varadaraj, Carina Magdaleno and Karthikeyan Mythreye, date: 08/20/2018, view: 28, Q&A: 0
    [Abstract] Fibronectin (FN) is an extracellular matrix protein that is secreted by many cell types and binds predominantly to the cell surface receptor Integrin α5β1. Integrin α5β1 binding initiates the step-wise assembly of FN into fibrils, a process called fibrillogenesis. We and several others have demonstrated critical effects of fibrillogenesis on cell ...
    Isolation of LYVE-1+ Endothelial Cells from Mouse Embryos
    Author:  Patrick Crosswhite, date: 08/05/2018, view: 118, Q&A: 0
    [Abstract] Lymphatic vessel endothelial hyaluronan receptor 1, or LYVE-1, is a type 1 integral membrane glycoprotein expressed by lymphatic endothelial cells (LECs). LYVE-1 is commonly used as a biological marker to visually distinguish developing lymphatic vessels from blood endothelial cells (arteries or veins). As our understanding of lymphatic biology is ...
    Hypoxia Reporter Element Assay
    Authors:  Daelynn R Buelow and Sharyn D. D Baker, date: 08/05/2018, view: 117, Q&A: 0
    [Abstract] Hypoxia is a condition in which there is a decrease in oxygen supply to the cellular environment. Changes to cellular oxygen levels can lead to transcriptional changes of oxygen-regulated genes. Reporter assays are used to study gene expression alteration and modifications in response to environmental changes. Dual-reporter assays allow the ...
    Quantitative Electron Microscopic Assay Using Random Sampling from Single Sections to Test Plastic Synaptic Changes in Hippocampus
    Authors:  G. Mark Marcello, Lilla E. Szabó, Peter Sotonyi and Bence Rácz, date: 08/05/2018, view: 135, Q&A: 0
    [Abstract] Studies over several decades on the organization of the CA1 hippocampus–a particularly favorable model for learning, memory and certain forms of cognition–have shown that the synaptic network in this brain region is plastic (Fortin et al., 2012). Recent evidence suggests that a number of environmental and endogenous stimuli may have a ...