tetano
Editor, Senior Moderator
Methods Mol Biol. 2020;2098:141-147. doi: 10.1007/978-1-0716-0207-2_9. [h=1]Influenza A Virus-Infected Lung Epithelial Cell Co-Culture with Human Peripheral Blood Mononuclear Cells.[/h]
Loh L[SUP]1,[/SUP][SUP]2[/SUP], Koutsakos M[SUP]3[/SUP], Kedzierska K[SUP]3[/SUP], Hinks TSC[SUP]4[/SUP].
[h=3]Author information[/h] 1 Department of Microbiology and Immunology, The Peter Doherty Institute for Infection and Immunity, The University of Melbourne, Parkville, Australia. lohl@unimelb.edu.au. 2 Department of Immunology and Microbiology, University of Colorado Anschutz Medical Campus, Aurora, CO, USA. lohl@unimelb.edu.au. 3 Department of Microbiology and Immunology, The Peter Doherty Institute for Infection and Immunity, The University of Melbourne, Parkville, Australia. 4 Respiratory Medicine Unit, Nuffield Department of Medicine Experimental Medicine, University of Oxford, Oxfordshire, UK.
[h=3]Abstract[/h] Sensing of influenza A virus (IAV) infection by pattern recognition receptors can occur by either direct infection of lung epithelial cells or uptake of virus-infected cells by innate cells such as dendritic cells/monocytes. This triggers a series of downstream events including activation of the inflammasome, the production of cytokines, chemokines, and the upregulation of stress-induced ligands that can lead to the activation of innate cells. These cells include innate lymphocytes such as MAIT, NKT, NK, and γδ T cells. Here we describe a method used to allow activation of human innate lymphocytes in co-culture with an IAV-infected human lung epithelial cell line (A549) to measure ex vivo effector functions (TNF and IFNγ) in a mixed culture environment. We describe (1) infection of the human lung epithelial cell line, (2) co-culture with PBMC, and (3) measurement of activation using intracellular cytokine staining.
[h=4]KEYWORDS:[/h] Epithelial cell; Flow cytometry; Human; Infection; MAIT cell; Tetramer; Virus
PMID: 31792820 DOI: 10.1007/978-1-0716-0207-2_9
Loh L[SUP]1,[/SUP][SUP]2[/SUP], Koutsakos M[SUP]3[/SUP], Kedzierska K[SUP]3[/SUP], Hinks TSC[SUP]4[/SUP].
[h=3]Author information[/h] 1 Department of Microbiology and Immunology, The Peter Doherty Institute for Infection and Immunity, The University of Melbourne, Parkville, Australia. lohl@unimelb.edu.au. 2 Department of Immunology and Microbiology, University of Colorado Anschutz Medical Campus, Aurora, CO, USA. lohl@unimelb.edu.au. 3 Department of Microbiology and Immunology, The Peter Doherty Institute for Infection and Immunity, The University of Melbourne, Parkville, Australia. 4 Respiratory Medicine Unit, Nuffield Department of Medicine Experimental Medicine, University of Oxford, Oxfordshire, UK.
[h=3]Abstract[/h] Sensing of influenza A virus (IAV) infection by pattern recognition receptors can occur by either direct infection of lung epithelial cells or uptake of virus-infected cells by innate cells such as dendritic cells/monocytes. This triggers a series of downstream events including activation of the inflammasome, the production of cytokines, chemokines, and the upregulation of stress-induced ligands that can lead to the activation of innate cells. These cells include innate lymphocytes such as MAIT, NKT, NK, and γδ T cells. Here we describe a method used to allow activation of human innate lymphocytes in co-culture with an IAV-infected human lung epithelial cell line (A549) to measure ex vivo effector functions (TNF and IFNγ) in a mixed culture environment. We describe (1) infection of the human lung epithelial cell line, (2) co-culture with PBMC, and (3) measurement of activation using intracellular cytokine staining.
[h=4]KEYWORDS:[/h] Epithelial cell; Flow cytometry; Human; Infection; MAIT cell; Tetramer; Virus
PMID: 31792820 DOI: 10.1007/978-1-0716-0207-2_9