title: T Cells Capture Bacteria by Transinfection from Dendritic Cells.
abstract: Recently, we have shown, contrary to what is described, that CD4(+) T cells, the paradigm of adaptive immune cells, capture bacteria from infected dendritic cells (DCs) by a process called transinfection. Here, we describe the analysis of the transinfection process, which occurs during the course of antigen presentation. This process was unveiled by using CD4(+) T cells from transgenic OTII mice, which bear a T cell receptor (TCR) specific for a peptide of ovoalbumin (OVAp), which therefore can form stable immune complexes with infected dendritic cells loaded with this specific OVAp. The dynamics of green fluorescent protein (GFP)-expressing bacteria during DC-T cell transmission can be monitored by live-cell imaging and the quantification of bacterial transinfection can be performed by flow cytometry. In addition, transinfection can be quantified by a more sensitive method based in the use of gentamicin, a non-permeable aminoglycoside antibiotic killing extracellular bacteria but not intracellular ones. This classical method has been used previously in microbiology to study the efficiency of bacterial infections. We hereby explain the protocol of the complete process, from the isolation of the primary cells to the quantification of transinfection.
authors: Cellular and Molecular Biology, Spanish National Research Council (CSIC), Spanish National Biotechnology Centre (CNB-CSIC); acruz@cnb.csic.es.
authors: Aranzazu
authors: A
authors: Cruz-Adalia
authors: Cellular and Molecular Biology, Spanish National Research Council (CSIC), Spanish National Biotechnology Centre (CNB-CSIC); Hospital de Santa Cristina, Healthcare Research Institute Princesa Hospital (IIS-Princesa).
authors: Guillermo
authors: G
authors: Ramírez-Santiago
authors: Cellular and Molecular Biology, Spanish National Research Council (CSIC), Spanish National Biotechnology Centre (CNB-CSIC); Hospital de Santa Cristina, Healthcare Research Institute Princesa Hospital (IIS-Princesa).
authors: Mónica
authors: M
authors: Torres-Torresano
authors: Cellular and Molecular Biology, Spanish National Research Council (CSIC), Spanish National Biotechnology Centre (CNB-CSIC); Hospital de Santa Cristina, Healthcare Research Institute Princesa Hospital (IIS-Princesa).
authors: Raquel
authors: R
authors: Garcia-Ferreras
authors: Cellular and Molecular Biology, Spanish National Research Council (CSIC), Spanish National Biotechnology Centre (CNB-CSIC); eveiga@cnb.csic.es.
authors: Esteban
authors: E
authors: Veiga Chacón
mesh_terms: D000818:Animals; D017951:Antigen Presentation; D001419:Bacteria / Q000378:metabolism; D001424:Bacterial Infections / Q000276:immunology* / Q000382:microbiology*; D015496:CD4-Positive T-Lymphocytes / Q000276:immunology* / Q000382:microbiology*; D003713:Dendritic Cells / Q000276:immunology* / Q000382:microbiology*; D005434:Flow Cytometry; D049452:Green Fluorescent Proteins / Q000096:biosynthesis; D051379:Mice; D008810:Mice, Inbred C57BL; D008822:Mice, Transgenic; D011948:Receptors, Antigen, T-Cell / Q000276:immunology
chemical_list: D011948:Receptors, Antigen, T-Cell; D049452:Green Fluorescent Proteins