Increase in viral antigen in individual polyoma-virus-infected mouse kidney cells as studied by quantiative immunofluorescence

Polyoma-virus infection of mouse kidney cells in vitro was studied on the single cell level by quantitative cytochemical techniques. Cells were fixed in acetone at different times after infection and reacted with fluorescein-conjugated mouse anti-polyoma-virus immune serum. Visual determinations were made as to whether the cells were fluorescent (“positive”) or not (“negative”). Visually the fluorescence was located in the nuclei in the “positive” cells. The fluorescence intensities of individual cell nuclei were then measured in a microspectrofluorimeter. Nuclei with considerable fluorescence intensity were noted to be visually “positive”. However, nuclei of cells which were usually judged to be “doubtful” could also be shown to have measurably greater fluorescence intensity than control non-infected cells. Increase in fluorescence values were noted 6-11 h after infection whereas hemagglutination tests did not become positive until 10-20 h later. With increasing time post infection, the fraction of fluorescence positive cells increased both as judged visually and as determined by the microfluorimeter. The variation in fluorescence intensities among different nuclei also increased and, 48-50 h after infection, the ratio between the highest and the lowest fluorescence values of positive cells was about 10:1. Higher fluorescence intensities were found in cytoplasms of cells with “positive” nuclei than in cells with “negative” nuclei. The cytoplasmic and nuclear fluorescence intensities correlated well in cells with “positive nuclei”. The polyoma-virus infection-induced stimulation of DNA synthesis was demonstrated by increased incorporation of ³H-thymidine and by increased amounts of DNA in the cells as measured by Feulgen microspectrophotometry. The amount of DNase-resistant DNA (i.e. viral DNA) correlated well with the intensity of immunofluorescence measured on the same individual nuclei of infected cells. This was taken as one indication that viral antigens can be measured on the single cell level by quantitative immunofluorimetry.