title: Methods suitable for high-throughput screening of siRNAs and other chemical compounds with the potential to inhibit rotavirus replication.
abstract: Rotaviruses are an important cause of severe gastroenteritis in children under two years of age. Two vaccines have become recently available, however, there are no specific pharmacological interventions of rotavirus disease. Recently, libraries of siRNAs or libraries of chemical compounds that can be tested for their ability to inhibit biological processes have been developed. To search these libraries for drugs or siRNAs that may prevent rotavirus replication it is necessary to have methods for high-throughput screening. In this study several methods to quantify rotavirus replication in cell culture were evaluated; the cell death and viral protein expression assays were compared, and an in-cell Western method based on infrared detection that allows the simultaneous quantification of viral antigen and total protein content in the same cell culture well was developed. This is an easy, inexpensive method for detection of viral replication, and it is compatible with high-throughput screening.
authors: Departamento de Genética del Desarrollo y Fisiología Molecular, Instituto de Biotecnología, UNAM, Avenida Universidad 2001, Colonia Chamilpa, Cuernavaca, Morelos 62210, Mexico. tdlopez@ibt.unam.mx
authors: Tomás
authors: T
authors: López
authors: Daniela
authors: D
authors: Silva-Ayala
authors: Susana
authors: S
authors: López
authors: Carlos F
authors: CF
authors: Arias
mesh_terms: D000818:Animals; D000956:Antigens, Viral / Q000096:biosynthesis; D000998:Antiviral Agents / Q000302:isolation & purification*; D001688:Biological Products / Q000302:isolation & purification*; D002470:Cell Survival; D002478:Cells, Cultured; D004353:Drug Evaluation, Preclinical / Q000379:methods*; D057166:High-Throughput Screening Assays / Q000379:methods*; D006801:Humans; D034741:RNA, Small Interfering / Q000302:isolation & purification*; D012401:Rotavirus / Q000187:drug effects*; D014779:Virus Replication / Q000187:drug effects*
chemical_list: D000956:Antigens, Viral; D000998:Antiviral Agents; D001688:Biological Products; D034741:RNA, Small Interfering