In situ ELISA for the evaluation of antiviral compounds effective against human cytomegalovirus

An in situ ELISA was developed as an improved procedure over the plaque reduction assay for antiviral susceptibility testing of HCMV. Unlike the plaque reduction assay, the ELISA can be completed at 4–5 days post-infection. The effective dose (ED 50) of ganciclovir (GCV), acyclovir (ACV), phosphonoacetic acid (PAA), or phosphonoformic acid (PFA), was determined using HCMV strain AD169. The resistance profiles of two laboratory-derived GCV-resistant mutants of HCMV strain AD169 and seven clinical isolates were determined using the ELISA. The ELISA results were confirmed by the plaque reduction assay. The ED 50 for GCV with the AD169 control ranged from 3.1 to 6.2 μM with a mean inhibitory concentration of 5.4 ± 1.4 μM. Six of the clinical isolates were susceptible to GCV (ED 50 = 3.1–6.2 μM). The seventh isolate had an ED 50 of 50 μM and was resistant to GCV. This ELISA assay is reproducible and relatively simple to perform. The ELISA endpoints are clearly determined and the assay works well with a variety of antiviral compounds.