Rapid detection of respiratory syncytial virus in nasopharyngeal secretions by enzyme-linked immunosorbent assay

Abstract

A new enzyme-linked immunosorbent assay (ELISA) respiratory syncytial virus antigen detection kit (Ortho Diagnostics, Inc., Raritan, N.J.) was compared with virus culture and with the indirect fluorescent antibody test (FAT) by using fresh nasal washings from children with suspected respiratory syncytial virus infection. Both uncentrifuged nasal washings and pellets from centrifuged split specimens were examined by ELISA. The ELISA was considered positive when the optical density was greater than the mean background optical density plus 0.15. Specimens positive by ELISA but negative by culture and FAT were reexamined in an ELISA blocking assay. Of 337 uncentrifuged specimens, 124 (37%) were positive by culture, 107 (32%) were positive by FAT, and 166 (49%) were positive by ELISA. Blocking assays showed that 21 of 30 (70%) of the seemingly false-positive ELISA tests were, in fact, true-positives and that the cultures and FATs were falsely negative. A patient specimen was considered positive when it was positive by virus culture, FAT, or blocking assay. The sensitivity, specificity, and positive predictive value of the ELISA test were 88, 94, and 95%, respectively. Centrifugation of nasal washings raised the sensitivity from 88 to 92% but reduced the specificity from 94 to 81%. We conclude that the Ortho ELISA test of uncentrifuged nasal washings is more sensitive than virus culture or indirect FAT and is highly specific.

Comparison of nasopharyngeal aspirate and nasopharyngeal swab specimens for respiratory syncytial virus diagnosis by cell culture, indirect immunofluorescence assay, and enzyme-linked immunosorbent assay

“…The sensitivity of the IFA procedure for RSV antigen detection was similar to that of ELISA in our study and those of others (9,12,13,15,16,22,24). For the IFA procedure, NPA specimens were better than NPS specimens owing to the greater number of cells collected by the former procedure.…”

“…Although the IFA technique is rapid, it is less readily adapted as a routine procedure because of difficulties with specimen preparation and reader subjectivity. Unfortunately, apparent false-positive results may occur when specimens are collected at a later stage of the disease (15) or if transport and virus culture are suboptimal. However, the IFA technique will benefit greatly from the use of monoclonal antibodies, which have shown results comparable to polyclonal reagents (2).…”

Detection, pathogenesis, and therapy of respiratory syncytial virus infections.

“…Hendry and colleagues (34) found that immunofluorescence was superior to ELISA when intact cells were used as the source of target antigen, but minimal differences were seen when whole secretions were used. In contrast, Lauer and colleagues (53) found sensitivities for ELISAs to be between 49 and 57% with uncentrifuged and centrifuged specimens, respectively. Sensitivities with immunofluorescence assays on the same specimens ranged from 32 to 47% in uncentrifuged and centrifuged specimens.…”

“…ELISA kits for the detection of RSV antigen in nasopharyngeal secretions are now commercially available, and several studies that evaluate their utility have appeared (9,53,82). Assays performed with the kits from Ortho Diagnostics and Abbott Laboratories have demonstrated a sensitivity (versus cell culture isolation) of 87 to 94% and specificity ranging from 72 to 95%.…”

“…Nasopharyngeal aspirates appear to be preferable to nasal swabs for obtaining suitable specimens for ELISAs. Centrifugation of the specimen before testing seems to increase the sensitivity somewhat, but at a drastic reduction in specificity in comparison to uncentrifuged specimens (53). The use of whole samples of secretions also seems to result in significantly greater sensitivities of the ELISA, in comparison to assays with washed cells as the target (34).…”