title: Measurement of mouse cytomegalovirus-induced interferon-β with immortalized luciferase reporter cells.
abstract: The production of cytokines is a crucial element of the host response to viral and bacterial infections. To follow these events in vivo, transgenic mice have become a valuable tool to study cytokine production through induction of reporter genes. We describe here the generation and immortalization of cells derived from transgenic reporter mice for development of a high-throughput assay system for virus- or bacteria-induced cytokine induction. As an example we describe mouse cytomegalovirus (MCMV) infection of immortalized fibroblasts derived from mice expressing the firefly luciferase reporter downstream of the IFN-β promoter. Common methods to determine IFN-β production, including ELISA, quantitative real-time PCR (qPCR), and transient reporter assays using plasmid-based reporter constructs, have disadvantages and limitations. Transient transfections influence type I IFN responses in most cell types, and IFN-β ELISA as well as qPCR are both laborious and expensive. The method presented here is highly sensitive as well as cost-effective, and allows monitoring of a robust and dose-dependent induction of IFN-β upon virus infection in cell lysates as well as living cells.
authors.affiliation: Helmholtz Centre for Infection Research, Braunschweig, Germany.
authors.forename: Evgenia
authors.initials: E
authors.lastname: Scheibe
authors.forename: Stefan
authors.initials: S
authors.lastname: Lienenklaus
authors.forename: Tobias
authors.initials: T
authors.lastname: May
authors.forename: Vladimir Gonçalves
authors.initials: VG
authors.lastname: Magalhães
authors.forename: Siegfried
authors.initials: S
authors.lastname: Weiss
authors.forename: Melanie M
authors.initials: MM
authors.lastname: Brinkmann
mesh_terms: D000818:Animals; D000952:Antigens, Polyomavirus Transforming / Q000235:genetics; D002461:Cell Line, Transformed; D015925:Cryopreservation / Q000379:methods; D005347:Fibroblasts / Q000378:metabolism / Q000821:virology; D015870:Gene Expression; D017930:Genes, Reporter; D005822:Genetic Vectors / Q000235:genetics; D057809:HEK293 Cells; D006801:Humans; D016899:Interferon-beta / Q000096:biosynthesis*; D016086:Lentivirus / Q000235:genetics; D008156:Luciferases / Q000235:genetics* / Q000378:metabolism; D051379:Mice; D018146:Muromegalovirus / Q000276:immunology*; D011401:Promoter Regions, Genetic; D014161:Transduction, Genetic
chemical_list: D000952:Antigens, Polyomavirus Transforming; D016899:Interferon-beta; D008156:Luciferases