Immunology

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    Protocols in Current Issue
    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: 842, 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 ...
    Evaluation of Cross-presentation in Bone Marrow-derived Dendritic Cells in vitro and Splenic Dendritic Cells ex vivo Using Antigen-coated Beads
    Authors:  Andrés Alloatti, Fiorella Kotsias, Eik Hoffmann and Sebastian Amigorena, date: 11/20/2016, view: 6017, Q&A: 0
    [Abstract] Antigen presentation by MHC class I molecules, also referred to as cross-presentation, elicits cytotoxic immune responses. In particular, dendritic cells (DC) are the most proficient cross-presenting cells, since they have developed unique means to control phagocytic and degradative pathways.

    This protocol allows the evaluation of ...
    Analysis of Phagosomal Antigen Degradation by Flow Organellocytometry
    [Abstract] Professional phagocytes internalize self and non-self particles by phagocytosis to initiate innate immune responses. After internalization, the formed phagosome matures through fusion and fission events with endosomes and lysosomes to obtain a more acidic, oxidative and hydrolytic environment for the degradation of its cargo. Interestingly, ...
    Phagocytosis Assay to Measure Uptake of Necroptotic Cancer Cells by BMDCs
    Authors:  Tania Løve Aaes, Dmitri V. Krysko and Peter Vandenabeele, date: 11/05/2016, view: 6926, Q&A: 0
    [Abstract] This protocol is a flow cytometry-based method to measure the phagocytosis efficiency of necroptotic target cells by bone marrow-derived dendritic cells (BMDCs) in vitro (Aaes et al., 2016). The method is a slightly modified and updated version of the protocols used in previously published papers (Krysko et al., 2006; ...
    Skin TRITC Painting to Track Dendritic Cells Migrating to the Lymph Nodes
    Authors:  Jyh Liang Hor and Scott N. Mueller, date: 09/20/2016, view: 5425, Q&A: 0
    [Abstract] Our protocol describes a simple method that allows tracking of dendritic cells (DC) migration from the flank skin to draining lymph nodes (LN) using a red fluorescent dye tetramethylrhodamine-5-isothiocyanate (TRITC). TRITC is a photostable dye that readily labels cells including DC in the skin and can survive repeated exposure of two-photon laser ...
    In vivo DCs Depletion with Diphtheria Toxin and MARCO+/MOMA1+ Cells Depletion with Clodronate Liposomes in B6.CD11c-DTR Mice
    [Abstract] To evaluate precisely the relative roles of different splenic phagocytic cells during an immune response, efficient methods for the depletion of specific populations are needed. Here, we describe the protocols for the depletion of splenic dendritic cells (DCs) by human diphtheria toxin (DTx) treatment in target mice (which express the human DTx ...
    Cell Tracer Violet and CellTracker Red CMTPX Staining of Purified Mature Plasmodium-infected Red Blood Cells
    [Abstract] Efficient staining methods to identify Plasmodium-infected red blood cells (iRBCs) are crucial to discriminate precisely the immune cells responsible for their elimination from circulation. Here, we describe the protocol for the purification of iRBCs and their subsequent staining with the vital dyes Cell Tracer Violet (CTV) or CellTracker ...
    Analysis of T Cell Proliferating and Polarizing Potential of Murine Dendritic Cells in Allogeneic-mixed Leukocyte Reaction
    Authors:  Pawan Kumar and Sangeeta Bhaskar, date: 03/05/2016, view: 6316, Q&A: 0
    [Abstract] Dendritic cells (DCs) play a critical role in mounting the T cell response against different infectious agents. Nature and intensity of the induced T cell responses are defined by activation status of DCs. It is generally accepted that IL-12, IL-4/IL-5 and IL-23 producing DCs induce TH1, TH2 and TH17 type of immune ...
    Generation and Maturation of Human Monocyte-derived DCs
    Authors:  Michela Spadaro, Monica Montone and Federica Cavallo, date: 08/05/2014, view: 17803, Q&A: 0
    [Abstract] Dendritic cells (DC) are antigen-presenting cells, which play a critical role in the regulation of the adaptive immune response. They act as a bridge between the innate and the adaptive immune systems. An approach to study their function and potentiality is to generate DC-like cells by culturing CD14+ monocyte-enriched peripheral blood ...
    Assessment of Human Dendritic Cell Antigen Uptake by Flow Cytometry
    Authors:  Ana Luque, Sonia Cárdenas-Brito, Rut Olivar and Josep M. Aran, date: 11/20/2013, view: 11583, Q&A: 0
    [Abstract] Antigen uptake by dendritic cells is the first key step towards induction of antigen-specific T-cell responses. This flow cytometry-based protocol describes the analysis of dendritic cell uptake of soluble antigens through two different mechanisms: non-specific macropinocytosis (using Lucifer Yelloy CH), and receptor-mediated endocytosis (using DQ ...



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