Protocols in Current Issue
    Characterization of Mouse Adult Testicular Macrophage Populations by Immunofluorescence Imaging and Flow Cytometry
    Authors:  Noushin Mossadegh-Keller and Michael H. Sieweke, date: 03/05/2019, view: 673, Q&A: 0
    [Abstract] Testicular macrophages (tMΦ) are the most abundant immune cells residing in the testis, an immune-privileged organ. TMΦ are known to exhibit different functions, such as protecting spermatozoa from auto-immune attack by producing immunosuppressive cytokines and trophic roles in supporting spermatogenesis and male sex hormone production. They also ...
    Immunohistochemical Staining of TLR4 in Human Skeletal Muscle Samples
    Authors:  Daniela Verzola, Samantha Milanesi, Francesca Costigliolo and Giacomo Garibotto, date: 01/20/2019, view: 818, Q&A: 0
    [Abstract] Growing evidence suggests the involvement of TLR4, a receptor in the innate immune system, in muscle loss in uremia. Recently, we have evaluated TLR4 in human skeletal muscle from chronic kidney disease patients, by immunohistochemistry and image analysis. Unlike the commonly-used Western blot method, immunohistochemistry allows for the ...
    Quantification of Extracellular Double-stranded RNA Uptake and Subcellular Localization Using Flow Cytometry and Confocal Microscopy
    Authors:  Tan A Nguyen, Lachlan Whitehead and Ken C Pang, date: 06/20/2018, view: 1881, Q&A: 0
    [Abstract] Double-stranded RNA is a potent pathogen-associated molecular pattern (PAMP) produced as a by-product of viral replication and a well-known hallmark of viral infection. Viral dsRNAs can be released from infected cells into the extracellular space and internalized by neighboring cells via endocytosis. Mammals possess multiple pattern recognition ...
    Proximal Ligation Assay (PLA) on Lung Tissue and Cultured Macrophages to Demonstrate Protein-protein Interaction
    Authors:  Roberto Mendez and Santanu Banerjee, date: 11/05/2017, view: 3976, Q&A: 0
    [Abstract] In this protocol, we describe proximal ligation assay (PLA), an antibody-based detection method for protein-protein interaction. This method relies on specific binding of individual primary antibodies to the two putative interacting proteins. The primary antibodies need to have different hosts. The secondary antibodies against the two hosts have ...
    In vitro Detection of Neutrophil Traps and Post-attack Cell Wall Changes in Candida Hyphae
    Authors:  Alex Hopke and Robert T. Wheeler, date: 04/05/2017, view: 4203, Q&A: 0
    [Abstract] In this protocol we describe how to visualize neutrophil extracellular traps (NETs) and fungal cell wall changes in the context of the coculture of mouse neutrophils with fungal hyphae of Candida albicans. These protocols are easily adjusted to test a wide array of hypotheses related to the impact of immune cells on fungi and the cell ...
    Ear Inflammation and Whole-mount Ear Staining
    Authors:  Jia Tong Loh, Merry Gunawan and I-hsin Su, date: 10/20/2016, view: 5318, Q&A: 0
    [Abstract] The recruitment of circulating neutrophils from the bloodstream to the site of inflammation represents one of the earliest events during an innate immune response. During this response, neutrophils tether and roll along the vessel walls before transmigrating across the endothelium into the interstitial space to exert their functions. Here, we ...
    Imaging Thick Lymph Node Tissue Sections
    Authors:  Jyh Liang Hor and Scott N. Mueller, date: 09/20/2016, view: 6570, Q&A: 0
    [Abstract] Our protocol describes a simple procedure for imaging thick lymph node sections by 2-photon microscopy. Lymph nodes are sectioned using a vibratome (vibrating microtome) to produce slices of tissue that can then be stained with fluorescently labeled antibodies. The thick tissue sections (150-200 μm depth) allow for the detection of cell clustering ...
    Confocal Imaging of Myeloid Cells in the Corneal Stroma of Live Mice
    Authors:  Matilda F. Chan and Zena Werb, date: 07/05/2015, view: 7035, Q&A: 0
    [Abstract] The accessibility and transparency of the cornea makes it a good tissue model for monitoring immunological responses using in vivo real time imaging analysis (Lee et al., 2010; Tan et al., 2013). These corneal qualities have also allowed for high-resolution in vivo imaging of non-ocular tissue transplanted into ...

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