Colorimetric Lactate Dehydrogenase (LDH) Assay for Evaluation of Antiviral Activity against Bovine Viral Diarrhoea Virus (BVDV) in Vitro

A rapid and sensitive screening assay has been established for in vitro evaluation of antiviral compounds against bovine viral diarrhoea virus (BVDV), which is widely used as a surrogate for hepatitis C virus (HCV). The procedure is based on photospectrometrical assessment for the viability of virus-infected cells via extracellular leakage of lactic dehydrogenase (LDH). The level of LDH in culture supernatants of BVDV-infected Madin-Darby bovine kidney (MDBK) cells was significantly higher than those of mock-infected MDBK cells. Under optimized assay conditions, the LDH level was found to correlate well with the degree of viral replication. When the 50% effective concentrations (EC50s) of ribavirin, cyclosporine A and human interferon-alpha for BVDV replication were determined by the established LDH method and compared with those obtained by a conventional tetrazolium colorimetric (MTT) method, there was a complete correlation in EC50s between the two methods. Furthermore, a much higher ratio of background activity (noise) to sample activity (signal) could be achieved with the LDH method than with the MTT method, indicating that the present LDH assay permits a sensitive, rapid and reliable screening of compounds for their anti-BVDV activity and may be useful for the discovery of novel anti-HCV agents.