title: Reverse transfected cell microarrays in infectious disease research.
abstract: Several human pathogenic viruses encode large genomes with often more than 100 genes. Viral pathogenicity is determined by carefully orchestrated co-operative activities of several different viral genes which trigger the phenotypic functions of the infected cells. Systematic analyses of these complex interactions require high-throughput transfection technology. Here we have provided a laboratory manual for the reverse transfected cell microarray (RTCM; alternative name: cell chip) as a high-throughput transfection procedure, which has been successfully applied for the systematic analyses of single and combination effects of genes encoded by the human herpesvirus-8 on the NF-kappaB signal transduction pathway. In order to quantitatively determine the effects of viral genes in transfected cells, protocols for the use of GFP as an indicator gene and for indirect immunofluorescence staining of cellular target proteins have been included. RTCM provides a useful methodological approach to investigate systematically combination effects of viral genes on cellular functions.
authors.affiliation: Division of Molecular and Experimental Surgery, Department of Surgery, University Medical Center Erlangen, Erlangen, Germany. andreas.konrad@uk-erlangen.de
authors.forename: Andreas
authors.initials: A
authors.lastname: Konrad
authors.forename: Ramona
authors.initials: R
authors.lastname: Jochmann
authors.forename: Elisabeth
authors.initials: E
authors.lastname: Kuhn
authors.forename: Elisabeth
authors.initials: E
authors.lastname: Naschberger
authors.forename: Priya
authors.initials: P
authors.lastname: Chudasama
authors.forename: Michael
authors.initials: M
authors.lastname: Stürzl
mesh_terms: D002461:Cell Line, Transformed; D003141:Communicable Diseases / Q000821:virology*; D019084:Fluorescent Antibody Technique, Indirect; D005814:Genes, Viral; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D057809:HEK293 Cells; D019288:Herpesvirus 8, Human / Q000235:genetics / Q000378:metabolism; D006801:Humans; D016328:NF-kappa B / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction / Q000235:genetics; D046888:Tissue Array Analysis / Q000379:methods*; D051996:Transcription Factor RelA / Q000378:metabolism; D014162:Transfection
chemical_list: D016328:NF-kappa B; D051996:Transcription Factor RelA; D049452:Green Fluorescent Proteins