What are RCAs?
One concern when working with adenoviral vectors is the rare occurrence of replication competent adenoviruses (RCAs) in a population of replication-deficient viruses. RCAs can emerge as a result of the rare double crossover through overlapping sequences present in the recombinant adenovirus and the genome of HEK293 cells. This event results in the replacement of the transgene by E1 region. Once this happens, the adenovirus could replicate, without the need of a complementing cell line. To detect RCA, non-complementary cells, such as A549 cells, are incubated with the viral stocks (1E4-1E6 PFU) and monitored for cytopathic effects (CPE) and/ or plaque formation.
