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 (ED50) 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 ED50 for GCV with the AD169 control ranged from 3.1 to 6.2 microM with a mean inhibitory concentration of 5.4 +/- 1.4 microM. Six of the clinical isolates were susceptible to GCV (ED50 = 3.1-6.2 microM). The seventh isolate had an ED50 of 50 microM 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.
Tatarowicz
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Department of Microbiology, Loyola University Medical Center, Maywood, Illinois 60153.
Lurain
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Thompson
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D000956:Antigens, Viral / Q000032:analysis; D000998:Antiviral Agents / Q000494:pharmacology*; D002478:Cells, Cultured; D003587:Cytomegalovirus / Q000187:drug effects*; D004353:Drug Evaluation, Preclinical / Q000379:methods; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D006801:Humans; D008826:Microbial Sensitivity Tests / Q000379:methods; D010948:Viral Plaque Assay
D000956:Antigens, Viral; D000998:Antiviral Agents