title: An improved protocol for measuring cytotoxic T cell activity in anatomic compartments with low cell numbers.
abstract: The study of target cell lysis and cytokine production are valuable tools to characterize antigen-specific T and NK cell function during virus infections. After localized infections in compartments such as the lung or the brain, however, cell numbers isolated from these organs are too low to perform standard assays with individual mice. Here, we report a few simple modifications of the classical 51Cr release assay allowing reduction of the number of required effector cells by a factor of 10 without loosing sensitivity or specificity. Using not more than 4x10(5) effector cells, we were able to study ex vivo virus-specific CTL or NK activity from the lungs of individual mice after infection with respiratory syncytial virus (RSV) and from the brains of mice infected with Borna disease virus (BDV). Flow cytometric analysis of interferon-gamma production by virus-specific T cells including appropriate controls was achieved with as few as 10(5) effector cells.
authors.affiliation: Children's Hospital, University of Freiburg, Mathildenstrasse 1, D-79106, Freiburg, Germany.
authors.forename: T
authors.initials: T
authors.lastname: Ostler
authors.forename: K
authors.initials: K
authors.lastname: Schamel
authors.forename: T
authors.initials: T
authors.lastname: Hussell
authors.forename: P
authors.initials: P
authors.lastname: Openshaw
authors.forename: J
authors.initials: J
authors.lastname: Hausmann
authors.forename: S
authors.initials: S
authors.lastname: Ehl
mesh_terms: D000818:Animals; D001890:Borna Disease / Q000276:immunology / Q000473:pathology; D001921:Brain / Q000166:cytology / Q000276:immunology; D001992:Bronchoalveolar Lavage Fluid / Q000166:cytology; D002460:Cell Line; D003601:Cytotoxicity Tests, Immunologic / Q000379:methods*; D005434:Flow Cytometry; D066298:In Vitro Techniques; D007694:Killer Cells, Natural / Q000276:immunology; D008168:Lung / Q000166:cytology / Q000276:immunology; D018655:Lymphocyte Count; D008216:Lymphocytic Choriomeningitis / Q000276:immunology / Q000473:pathology; D051379:Mice; D008807:Mice, Inbred BALB C; D008810:Mice, Inbred C57BL; D009928:Organ Specificity; D018357:Respiratory Syncytial Virus Infections / Q000276:immunology / Q000473:pathology; D013602:T-Lymphocytes, Cytotoxic / Q000276:immunology*