Detection of respiratory syncytial virus and rotavirus by enhanced chemiluminescence enzyme-linked immunosorbent assay

Abstract

The horseradish peroxidase-catalyzed oxidation of luminol was used in a chemiluminescence avidin-biotin enzyme-linked immunosorbent assay (CL-ABE) to detect respiratory syncytial virus (RSV) and rotavirus (ROV). This CL-ABE was carried out in a new apparatus constructed for the photographic registration of the light emission produced. When measured in a spectrophotometer, the light emission produced by the oxidation of luminol showed a peak emission at 425 nm. The chemiluminescence output reached maximum within 1 min after the initiation of luminol oxidation and diminished to one-half of maximum emission within 8 min. When titrations of purified ROV by avidin-biotin enzyme-linked immunosorbent assay (ABE) were monitored by CL-ABE and by conventional orthophenylenediamine (OPD) staining (OPD-ABE), the detection limits were 0.01 and 0.04 ng of ROV protein, respectively. Similar titrations of purified RSV gave detection limits of 0.2 and 0.8 ng of RSV protein by CL-ABE and OPD-ABE, respectively. When 26 RSV-positive samples of nasopharyngeal secretions (NPS) were titrated by CL-ABE and by OPD-ABE, the mean titers obtained were 737 and 254, respectively. When 19 ROV-positive fecal samples were titrated by CL-ABE and by OPD-ABE, the mean titers obtained were 82,000 and 29,000, respectively. When samples of NPS from 123 infants and children with acute lower respiratory disease were tested for the presence of RSV, 31 NPS samples were RSV positive by CL-ABE, and 29 of these 31 NPS samples were RSV positive by OPD-ABE.