Development and evaluation of an automatable focus reduction neutralisation test for the detection of measles virus antibodies using imaging analysis

Abstract

A plaque reduction neutralisation test (PRNT) is still regarded as the gold standard for the investigation of anti-measles immunity. In this study, an alternative simplified automatable focus reduction neutralisation test (AFRNT) based on the classical PRNT was developed. The AFRNT uses the conventional Edmonston strain of measles, immunoperoxidase staining with monoclonal antibodies, and automated plaque counts performed with AID ViruSpot software. The assay is performed in 96-well plates, requires 2 days, and is fully automatable. The AFRNT was evaluated in comparison with PRNT and Enzygnost anti-measles enzyme immunoassay (EIA). A total of 130 samples, which included two available WHO international anti-measles standards, sera from 90 patients, and 38 different lots of immunoglobulin products, were tested. Overall, good agreement was observed between EIA and both neutralisation tests; however, the EIA values for the immunoglobulin products and international standards were slightly but significantly higher than those of the neutralisation tests. The Bland–Altman analysis showed excellent agreement between AFRNT and PRNT. AFRNT is a fully automatable high-throughput neutralisation assay, which can be performed with measles and other types of viruses, including wild-type strains. It is perfectly suited for epidemiological and vaccine studies.