Detection of serotypic variants in viral preparations: sensitivity of a procedure for selection

The efficiency of an in vitro method (the breakthrough neutralization procedure) for selecting serotypic variants from preparations of human rhinovirus (HRV) 17 and a temperature sensitive strain (Ts-1) of HRV-2 was examined. Viruses were plaqued in the presence of homologous polyclonal antisera, and plaques which escaped neutralization were isolated. For control purposes, isolates were obtained by plaquing in the absence of antisera. Any clone that consistently (minimum of 2 tests) yielded a 4-fold or greater difference in serum neutralization titer compared to the parent virus when tested with antiserum to the parent, was considered a serotypic variant. The efficiency of the breakthrough neutralization procedure was calculated using varying mixtures of 2 related rhinovirus serotypes. Using this method, the ability to isolate serotypic subpopulations from a mixture was enhanced at least 7000-fold. This procedure allows the rapid isolation of variants which differ from the parent virus by as little as 4-fold in a serum neutralization test. Hence, viral preparations used to prepare reference grade reagents such as seed stocks and polyclonal and monoclonal antisera, can be tested for specificity prior to use.