The screening of a large number of potential agents (radiation and photochemicals) for the inactivation of viruses which contaminate transfusable blood products necessitated the development of a rapid method for assaying viral activity. Classical assays of the cytopathic effect of such marker viruses as herpes simplex virus and poliovirus may require up to several weeks. Two bacteriophage, T2 and PM2 (lipid-enveloped), were chosen as additional viruses for our studies because they are readily available and well-described in the literature. A microtiter plate assay was developed which measures viability of the phage in a suspension of the host bacterium based on the optical density (turbidity) of the medium at 405nm. Our collaborators In DPQC obtained T2 and PM2 phage and grew the required host bacteria, E. coli and Alteromonas espejiana, respectively. Growth curves were obtained by three methods: plaque assay on agar plates, and turbidity measurement of liquid suspensions in both culture tubes and microtiter plates. For the latter, we ran up to 12 replicates of each concentration point over a dilution range of 6 logs. The microtiter assay correlated well with the other two methods and end-point tiers were obtained at 24h for T2 and at 16h for PM2. The microtiter data permitted analysis of variance and was directly recorded into computer format. Computer programs were developed to aid in data analysis. This new technique has been successfully applied to the viral Inactivation project and is described in a manuscript that is being prepared for publication.
