ASSAY OF VARIOLA VIRUS BY THE FLUORESCENT CELL-COUNTING TECHNIQUES
ARMY BIOLOGICAL LABS FREDERICK MD
Pagination or Media Count:
A quantitative assay for infective variola virus particles was developed that is based on the enumeeration of cells containing fluorescent viral antigen after infection of McCoy cell monolayers. The direct fluorescent antibody technique was employed to stain cells. The efficiency of virus adsorption was markedly enhanced by centrifugation of virus inoculum onto McCoy cell monolayers at 500 x g for 15 minutes. By this procedure, a proportionality was obtained between the number of fluorescent cells and volume of inoculum. Observations on the sequential development of viral antigen within cells and counts of fluorescent cells, showed the optimal time for the enumeration of fluorescent cells was after an incubation period of 16 to 20 hours. A linear function existed between virus concentration and cell infecting units. The assay was shown to be highly sensitive, precise, and reproducible. Fluorescent cells were randomly distributed in infected coverslip cell monolayyers. The demonstration that the concentration of variola virus in aerosols can be determined within 24 hours exemplifies the rapidity and applicability of the fluorescent cell-countingg technique.