Guidelines for mouse and human DC generation
European Journal of Immunology, vol. 53, pp. e2249816
Abstract
Abstract This article is part of the Dendritic Cell Guidelines article series, which provides a collection of state‐of‐the‐art protocols for the preparation, phenotype analysis by flow cytometry, generation, fluorescence microscopy, and functional characterization of mouse and human dendritic cells (DC) from lymphoid organs and various non‐lymphoid tissues. This article provides protocols with top ticks and pitfalls for preparation and successful generation of mouse and human DC from different cellular sources, such as murine BM and HoxB8 cells, as well as human CD34 + cells from cord blood, BM, and peripheral blood or peripheral blood monocytes. We describe murine cDC1, cDC2, and pDC generation with Flt3L and the generation of BM‐derived DC with GM‐CSF. Protocols for human DC generation focus on CD34 + cell culture on OP9 cell layers for cDC1, cDC2, cDC3, and pDC subset generation and DC generation from peripheral blood monocytes (MoDC). Additional protocols include enrichment of murine DC subsets, CRISPR/Cas9 editing, and clinical grade human DC generation. While all protocols were written by experienced scientists who routinely use them in their work, this article was also peer‐reviewed by leading experts and approved by all co‐authors, making it an essential resource for basic and clinical DC immunologists.
Authors 26
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Manfred Bernhard Lutz corresponding
Affiliation as printed
Institute for Virology and Immunobiology University of Würzburg Würzburg Germany
Section lead authors
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Düsseldorf University Hospital · Heinrich Heine University Düsseldorf
Affiliation as printed
Institute of Medical Microbiology and Hospital Hygiene University of Düsseldorf Düsseldorf Germany
Section lead authors
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The University of Melbourne · Walter and Eliza Hall Institute of Medical Research
Affiliation as printed
Department of Medical Biology University of Melbourne Parkville VIC 3052 Australia
Immunology Division The Walter and Eliza Hall Institute of Medical Research Parkville VIC 3052 Australia
Department of Medical Biology, University of Melbourne, Parkville, VIC, 3052 Australia
Immunology Division, The Walter and Eliza Hall Institute of Medical Research, Parkville, VIC, 3052 Australia
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German Cancer Research Center · University of Tübingen
Affiliation as printed
Dendritic Cells in Infection and Cancer (F171) German Cancer Research Center (DKFZ) Heidelberg Germany
Department of Internal Medicine II University of Tübingen Tübingen Germany
Dendritic Cells in Infection and Cancer (F171), German Cancer Research Center (DKFZ), Heidelberg, Germany
Section lead authors
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Johannes Gutenberg University Mainz · University Medical Center of the Johannes Gutenberg University Mainz
Affiliation as printed
Institute of Molecular Medicine University Medical Center of the Johannes Gutenberg‐University Mainz Mainz 55131 Germany
Institute of Molecular Medicine, University Medical Center of the Johannes Gutenberg-University Mainz, Mainz, 55131 Germany
Section lead authors
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Affiliation as printed
Translational and Clinical Research Institute Newcastle University Newcastle upon Tyne NE2 4HH United Kingdom
Section lead authors
Translational and Clinical Research Institute, Newcastle University, Newcastle upon Tyne, NE2 4HH United Kingdom
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Affiliation as printed
Institute for Virology and Immunobiology University of Würzburg Würzburg Germany
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Centre National de la Recherche Scientifique · Inserm · Aix-Marseille Université · Centre d’Immunologie de Marseille-Luminy
Affiliation as printed
CNRS, INSERM, Aix Marseille Univ, Centre d'Immunologie de Marseille‐Luminy Turing Center for Living Systems Marseille France
CNRS, INSERM, Aix Marseille Univ, Centre d'Immunologie de Marseille-Luminy, Turing Center for Living Systems, Marseille, France
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Friedrich-Alexander-Universität Erlangen-Nürnberg · Universitätsklinikum Erlangen · Comprehensive Cancer Center Erlangen
Affiliation as printed
Comprehensive Cancer Center Erlangen European Metropolitan Area of Nuremberg (CCC ER‐EMN) Östliche Stadtmauerstraße 30 91054 Erlangen Germany
Deutsches Zentrum Immuntherapie (DZI) Ulmenweg 18 91054 Erlangen Germany
RNA‐based Immunotherapy, Hautklinik, Universitätsklinikum Erlangen (UKER) Friedrich‐Alexander‐Universität (FAU) Erlangen‐Nürnberg Erlangen Germany
RNA‐based Immunotherapy, Hautklinik, Universitätsklinikum Erlangen (UKER) Friedrich‐Alexander‐Universität (FAU) Erlangen‐Nürnberg Erlangen Germany
Section lead authors
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Universitätsklinikum Erlangen · Comprehensive Cancer Center Erlangen
Affiliation as printed
Comprehensive Cancer Center Erlangen European Metropolitan Area of Nuremberg (CCC ER‐EMN) Östliche Stadtmauerstraße 30 91054 Erlangen Germany
Deutsches Zentrum Immuntherapie (DZI) Ulmenweg 18 91054 Erlangen Germany
Laboratory of Dendritic Cell Biology Department of Dermatology University Hospital Erlangen Hartmannstraße 14 D‐91052 Erlangen Germany
Medical Immunology Campus Erlangen (MICE) D‐91054 Erlangen Germany
Comprehensive Cancer Center Erlangen European Metropolitan Area of Nuremberg (CCC ER‐EMN) Östliche Stadtmauerstraße 30 91054 Erlangen Germany
Deutsches Zentrum Immuntherapie (DZI), Ulmenweg 18, 91054 Erlangen, Germany
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Radboud University Nijmegen · Radboud University Medical Center · Radboud Institute for Molecular Life Sciences
Affiliation as printed
Department of Tumor Immunology Radboud Institute for Molecular Life Sciences Radboudumc Nijmegen the Netherlands
Department of Tumor Immunology, Radboud Institute for Molecular Life Sciences, Radboudumc, Nijmegen, the Netherlands
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Johannes Gutenberg University Mainz · University Medical Center of the Johannes Gutenberg University Mainz
Affiliation as printed
Institute of Medical Microbiology and Hygiene University Medical Center of the Johannes Gutenberg‐University Mainz Mainz 55131 Germany
Institute of Medical Microbiology and Hygiene, University Medical Center of the Johannes Gutenberg-University Mainz, Mainz, 55131 Germany
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Johannes Gutenberg University Mainz · University Medical Center of the Johannes Gutenberg University Mainz
Affiliation as printed
Institute of Medical Microbiology and Hygiene University Medical Center of the Johannes Gutenberg‐University Mainz Mainz 55131 Germany
Institute of Medical Microbiology and Hygiene, University Medical Center of the Johannes Gutenberg-University Mainz, Mainz, 55131 Germany
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Affiliation as printed
Institute for Virology and Immunobiology University of Würzburg Würzburg Germany
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Institut für Immunologie · Ludwig-Maximilians-Universität München
Affiliation as printed
Institute for Immunology, Biomedical Center, Faculty of Medicine Ludwig‐Maximilians‐University Munich Planegg‐Martinsried Germany
Institute for Immunology, Biomedical Center, Faculty of Medicine, Ludwig-Maximilians-University Munich, Planegg-Martinsried, Germany
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Affiliation as printed
Laboratory of Dendritic Cell Biology Department of Dermatology University Hospital Erlangen Hartmannstraße 14 D‐91052 Erlangen Germany
Medical Immunology Campus Erlangen (MICE) D‐91054 Erlangen Germany
Laboratory of Dendritic Cell Biology Department of Dermatology University Hospital Erlangen Hartmannstraße 14 D‐91052 Erlangen Germany
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National Institutes of Health · National Cancer Institute · Center for Cancer Research
Affiliation as printed
Experimental Immunology Branch, Center for Cancer Research, National Cancer Institute National Institutes of Health Bethesda MD 20892 USA
Experimental Immunology Branch, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD, 20892 USA
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The University of Melbourne · Walter and Eliza Hall Institute of Medical Research
Affiliation as printed
Department of Medical Biology University of Melbourne Parkville VIC 3052 Australia
Immunology Division The Walter and Eliza Hall Institute of Medical Research Parkville VIC 3052 Australia
Department of Medical Biology, University of Melbourne, Parkville, VIC, 3052 Australia
Immunology Division, The Walter and Eliza Hall Institute of Medical Research, Parkville, VIC, 3052 Australia
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Düsseldorf University Hospital · Heinrich Heine University Düsseldorf
Affiliation as printed
Institute of Medical Microbiology and Hospital Hygiene University of Düsseldorf Düsseldorf Germany
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Radboud University Nijmegen · Radboud University Medical Center · Radboud Institute for Molecular Life Sciences
Affiliation as printed
Department of Tumor Immunology Radboud Institute for Molecular Life Sciences Radboudumc Nijmegen the Netherlands
Department of Tumor Immunology, Radboud Institute for Molecular Life Sciences, Radboudumc, Nijmegen, the Netherlands
Section lead authors
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Inserm · Université Paris Sciences et Lettres · Institut Curie
Affiliation as printed
Institut Curie PSL Research University INSERM, U932, 26 rue d'Ulm Paris 75005 France
Institut Curie, PSL Research University, INSERM, U932, 26 rue d'Ulm, Paris, 75005 France
Section lead authors
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Kristin Seré Aachen Institute for Biomedical Engineering Department of Cell Biology Institute for Biomedical Engineering Department of Cell Biology
Affiliation as printed
Helmholtz Institute for Biomedical Engineering, RWTH Aachen University Aachen Germany
Institute for Biomedical Engineering Department of Cell Biology RWTH Aachen University Medical School Aachen Germany
Helmholtz Institute for Biomedical Engineering, RWTH Aachen University, Aachen, Germany
Institute for Biomedical Engineering, Department of Cell Biology, RWTH Aachen University Medical School, Aachen, Germany
Section lead authors
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Johannes Gutenberg University Mainz · University Medical Center of the Johannes Gutenberg University Mainz
Affiliation as printed
Institute of Medical Microbiology and Hygiene University Medical Center of the Johannes Gutenberg‐University Mainz Mainz 55131 Germany
Institute of Medical Microbiology and Hygiene, University Medical Center of the Johannes Gutenberg-University Mainz, Mainz, 55131 Germany
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Eindhoven University of Technology
Affiliation as printed
Institute for Complex Molecular Systems (ICMS) Eindhoven University of Technology Eindhoven The Netherlands
Laboratory of Immunoengineering, Department of Biomedical Engineering Eindhoven University of Technology Eindhoven The Netherlands
Institute for Complex Molecular Systems (ICMS), Eindhoven University of Technology, Eindhoven, The Netherlands
Laboratory of Immunoengineering, Department of Biomedical Engineering, Eindhoven University of Technology, Eindhoven, The Netherlands
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Huaming Xu Aachen Institute for Biomedical Engineering Department of Cell Biology Institute for Biomedical Engineering Department of Cell Biology
Affiliation as printed
Helmholtz Institute for Biomedical Engineering, RWTH Aachen University Aachen Germany
Institute for Biomedical Engineering Department of Cell Biology RWTH Aachen University Medical School Aachen Germany
Helmholtz Institute for Biomedical Engineering, RWTH Aachen University, Aachen, Germany
Institute for Biomedical Engineering, Department of Cell Biology, RWTH Aachen University Medical School, Aachen, Germany
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Martin Zenke Aachen Institute for Biomedical Engineering Department of Cell Biology Institute for Biomedical Engineering Department of Cell Biology
Affiliation as printed
Helmholtz Institute for Biomedical Engineering, RWTH Aachen University Aachen Germany
Institute for Biomedical Engineering Department of Cell Biology RWTH Aachen University Medical School Aachen Germany
Helmholtz Institute for Biomedical Engineering, RWTH Aachen University, Aachen, Germany
Institute for Biomedical Engineering, Department of Cell Biology, RWTH Aachen University Medical School, Aachen, Germany
Section lead authors
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