Development of a rapid and sensitive quantitative assay for rotavirus based on flow cytometry

Abstract

A very sensitive and accurate flow cytometry (FC) based method have developed to quantitate rotavirus infection in MA104 cells. Confluent cell monolayers were infected with serial dilutions of rotavirus SA11. After infection, the cells were recovered with the aid of trypsin and then reacted with monoclonal antibody M60 (specific for the rotavirus outer capsid protein, VP7), followed by a second antibody (anti-mouse IgG-FITC). A FACScan FC was used to estimate the number of infected cells, as well as the level of infection. Viral infection was optimised by varying the concentration of trypsin used in the maintenance medium. The FC method enables many cells to be screened quickly for infectivity, and can detect low levels of virus. This method can be adapted to monitor the presence of other viruses in clinical and environmental samples without the need for prolonged periods of adaptation to growth in tissue culture.