title: RSV Growth and Quantification by Microtitration and qRT-PCR Assays.
abstract: Defective interfering viral particles have been reported as important determinants of the course of viral infection, and they can markedly temper the virulence of the infection. Here, we describe a simple method, based on limiting dilution, for the removal of defective interfering particles from RSV. This method results in a high-titer viral preparation from both HEp-2 and Vero cell lines. We evaluated two concentrations of sucrose to stabilize the virus preparation, and demonstrate that RSV is stable when prepared and stored in 25 % sucrose at -152 °C. In addition, this chapter describes some commonly used methods of RSV titration, detection using microtitration and quantitative real-time RT-PCR, and the use of immunostaining for antigenic characterization.
authors.affiliation: Gastroenteritis and Respiratory Viruses Laboratory Branch, Division of Viral Diseases, National Center for Immunization and Respiratory Diseases, Centers for Disease Control and Prevention (CDC), 1600 Clifton Road NE, Mailstop G-18, Atlanta, GA, 30333, USA.
authors.forename: Hayat
authors.initials: H
authors.lastname: Caidi
authors.affiliation: Gastroenteritis and Respiratory Viruses Laboratory Branch, Division of Viral Diseases, National Center for Immunization and Respiratory Diseases, Centers for Disease Control and Prevention (CDC), 1600 Clifton Road NE, Mailstop G-18, Atlanta, GA, 30333, USA.
authors.forename: Jennifer L
authors.initials: JL
authors.lastname: Harcourt
authors.affiliation: Division of Viral Diseases, National Center for Immunization and Respiratory Diseases, Centers for Disease Control and Prevention (CDC), 1600 Clifton Road NE, Mailstop A-34, Atlanta, GA, 30333, USA. loh5@cdc.gov.
authors.forename: Lia M
authors.initials: LM
authors.lastname: Haynes
mesh_terms: D000818:Animals; D045744:Cell Line, Tumor / Q000821:virology; D002522:Chlorocebus aethiops; D003673:Defective Viruses / Q000254:growth & development*; D006801:Humans; D060888:Real-Time Polymerase Chain Reaction; D018113:Respiratory Syncytial Virus, Human / Q000235:genetics / Q000254:growth & development*; D012680:Sensitivity and Specificity; D014709:Vero Cells / Q000821:virology; D019562:Viral Load; D014776:Virus Cultivation / Q000379:methods*
keywords: Immunostaining; Limiting dilution; Plaque assay; Respiratory syncytial virus; qRT-PCR