Particle counting

As we have discussed, not all viruses replicate well in cell culture, and even of those that do, not all virus particles within a sample are infectious. Indeed, in many cases, for every one infectious particle up to 100 or more noninfectious particles may be produced from an  infected cell. Loss of infectivity can be the result of many events such as incorrect virion assembly or genome packaging, the presence of a spontaneous mutation within the packaged virus genome or lack of a final virion maturation step. The total number of particles can only be determined with the aid of an electron microscope. The procedure relies  on  the  use  of reference particles, usually latex  beads of uniform diameter (latex and  virus particles are distinguishable when using negative staining with phosphotung- state), which are mixed  with  the  virus  at known concentration (i.e. a known number of reference particles per unit  volume). By counting the number of virions and the number of reference particles within a grid, it is a simple matter to determine the ratio of virions to reference particles and hence calculate the virus count. The ratio of total particles to infectious particles is termed the particle/infectivity ratio, which is important to know when, for example, monitoring virus purification, or determining the state or age of a virus suspension.