Adventitious virus assays are performed as part of raw materials testing, cell-line characterization, and lot-release testing of biologicals such as monoclonal antibodies, gene therapy vectors, recombinant proteins, and vaccines. The testing methods follow guidance provided in the 9 CFR (bovine and porcine raw materials testing, and certain vaccine products) or Points to Consider documents (cell line characterization and evaluation of the majority of biologicals). The methodologies used and the types of adventitious viruses detected during testing of the various types of samples are discussed in this paper. The detection of adventitious viruses is quite rare, especially during evaluation of cell banks and biologicals produced in human, mouse, or insect cell substrates. The most common detection scenarios include bovine viral diarrhoea virus in foetal bovine serum samples, porcine parvovirus in porcine substrates, and murine minute virus, REO virus, and Cache Valley virus in Chinese hamster cell-derived bulk harvests. The two last-named viral entities are believed to be introduced via bovine serum used during the manufacturing process (during scale-up or during the entire process). Knowledge of the types of agents being detected is useful in designing viral clearance methodologies for purification processes and in engineering manufacturing processes and facilities.