SUPERNATANT RESCUE ASSAY VS POLYMERASE CHAIN-REACTION FOR DETECTION OF WILD-TYPE ADENOVIRUS-CONTAMINATING RECOMBINANT ADENOVIRUS STOCKS

The certification of recombinant adenoviruses prepared for clinical us e requires the exclusion of contaminating, replication-competent adeno virus (wild type virus). Polymerase chain reaction (PCR)-based detecti on assays have been developed that detect the presence of viral sequen ces present only in wild type adenoviruses. As an alternative, this re port describes a novel bioassay, designated the 'supernatant rescue as say', that detected minimal amounts of wild type virus mixed with high numbers of recombinant adenoviruses. This assay is based on the obser vation that minimal numbers of wild type adenovirus can be rescued fro m the supernatants of cells exposed to high multiplicities of infectio n (mois) of recombinant virus mixed with a known, minimal number of wi ld type virions. This assay was highly reproducible and routinely dete cted the presence of a single wild type virus mixed within 10(9) recom binant viruses. Under the experimental conditions employed, the supern atant rescue assay was significantly more sensitive than all three dif ferent PCR-detection assays.