tailieunhanh - Báo cáo y học: " An RNA replication-center assay for high content image-based quantifications of human rhinovirus and coxsackievirus infections"

Tuyển tập báo cáo các nghiên cứu khoa học quốc tế ngành y học dành cho các bạn tham khảo đề tài: An RNA replication-center assay for high content image-based quantifications of human rhinovirus and coxsackievirus infections | Jurgeit et al. Virology Journal 2010 7 264 http content 7 1 264 VIROLOGY JOURNAL RESEARCH Open Access An RNA replication-center assay for high content image-based quantifications of human rhinovirus and coxsackievirus infections 1 3 1 1 4 Andreas Jurgeit Stefan Moese Pascal Roulin Alexander Dorsch Mark Lotzerich Wai-Ming Lee Urs F Greber1 Abstract Background Picornaviruses are common human and animal pathogens including polio and rhinoviruses of the enterovirus family and hepatits A or food-and-mouth disease viruses. There are no effective countermeasures against the vast majority of picornaviruses with the exception of polio and hepatitis A vaccines. Human rhinoviruses HRV are the most prevalent picornaviruses comprising more than one hundred serotypes. The existing and also emerging HRVs pose severe health risks for patients with asthma or chronic obstructive pulmonary disease. Here we developed a serotype-independent infection assay using a commercially available mouse monoclonal antibody mabJ2 detecting double-strand RNA. Results Immunocytochemical staining for RNA replication centers using mabJ2 identified cells that were infected with either HRV1A 2 14 16 37 or coxsackievirus CV B3 B4 or A21. MabJ2 labeled-cells were immunocytochemically positive for newly synthesized viral capsid proteins from HRV1A 14 16 37 or CVB3 4. We optimized the procedure for detection of virus replication in settings for high content screening with automated fluorescence microscopy and single cell analysis. Our data show that the infection signal was dependent on multiplicity time and temperature of infection and the mabJ2-positive cell numbers correlated with viral titres determined in single step growth curves. The mabJ2 infection assay was adapted to determine the efficacy of antiviral compounds and small interfering RNAs siRNAs blocking enterovirus infections. Conclusions We report a broadly applicable rapid protocol to measure infection of cultured cells .