Differential diagnosis of pandemic (H1N1) 2009 infection by detection of haemagglutinin with an enzyme-linked immunoassay

Abstract

A sensitive and convenient immunoassay that can directly differentiate pandemic (H1N1) 2009 (pH1N1) virus from seasonal influenza virus can play an important role in the clinic. In the presented study, a double-sandwich ELISA (pH1N1 ELISA), based on two monoclonal antibodies against haemagglutinin (HA) of the pH1N1 virus, was developed. After laboratory determination of the sensitivity and specificity characteristics, the performance of this assay was evaluated in a cohort of 904 patients with influenza-like illness. All seven strains of pH1N1 virus tested were positive by pH1N1 ELISA, with an average lower detection limit of 10(3.0 ± 0.4) tissue culture infective dose (TCID)(50) /mL (or 0.009 ± 0.005 HA titre). Cross-reaction of the assay with seasonal influenza virus and other common respiratory pathogens was rare. In pH1N1-infected patients, the sensitivity of the pH1N1 ELISA was 92.3% (84/91, 95% CI 84.8-96.9%), which is significantly higher than that of the BD Directigen EZ Flu A + B test (70.3%, p <0.01). The specificity of pH1N1 ELISA in seasonal influenza A patients was 100.0% (171/171, 95% CI 97.9-100.0%), similar to that in non-influenza A patients (640/642, 99.7%, 95% CI 98.9-100.0%). The positive predictive value for pH1N1 ELISA was 97.7% and the negative predictive value was 99.1% in this study population with a pH1N1 prevalence of 10.1%. In conclusion, detection of HA of pH1N1 virus by immunoassay appears to be a convenient and reliable method for the differential diagnosis of pH1N1 from other respiratory pathogens, including seasonal influenza virus.

Rapid identification of imported influenza viruses at Xiamen International Airport via an active surveillance program

“…This study showed that the Flu Dot-ELISA has comparatively high sensitivity in border screening for influenza infection (96.6, 95% CI 92.7e98.7). The higher sensitivity of the assay may be attributed to two aspects: first, the enzyme-induced signal amplification system used confers sensitivity at least ten times higher than that of traditional rapid assays based on the colloidal gold system [16]; and second, the nasopharyngeal specimens we tested were one of the most appropriate sample types for virus detection using rapid tests [17,18]. Nevertheless, six of the 176 positive influenza isolates were missed by the rapid test; further improvements in sensitivity would be desirable.…”

A sensitive and specific antigen detection assay for Middle East respiratory syndrome coronavirus

“…The capture ELISA for MERS-CoV-NP detection was developed as described previously with minor modifications. 21 Microplates (Sigma Aldrich, Saint Louis, MO, USA) were pre-coated with the first anti-MERS-CoV-rNP MAb 1F6 and incubated at 37 °C overnight with a blocking reagent (phosphate-buffered saline with 2% sucrose, 0.2% casein-Na, and 2% gelatin). Inactivated MERS-CoV, HCoV-229E, HCoV-OC43, two influenza virus A strains, one influenza virus B strain and one respiratory syncytial virus (RSV) strain, and purified rNPs of MERS-CoV, HCoV-229E, and HCoV-OC43 were diluted in phosphate-buffered saline with 2% skim milk.…”

Molecular Diagnostics as an Indispensable Tool for the Diagnosis of Infectious Diseases of Viral Origin and Global Impact

“…These tools are continuously improved, a double-sandwich ELISA (pH1N1 ELISA), based on two monoclonal antibodies against haemagglutinin (HA) of the pH1N1 virus has a sensitivity of 92.3% (84/91, 95% CI 84.8-96.9%), being significantly higher than that of the BD Directigen EZ Flu A+B test (70.3%, p <0.01). In addition, this assay can directly differentiate pandemic (H1N1) 2009 (pH1N1) virus from other respiratory pathogens, including seasonal influenza virus [56]. A hemagglutination inhibition assay is an extremely reliable tool for typing, subtyping and analyze the antigenic characteristics of influenza viral isolates if the reference antisera used contain antibodies to currently circulating viruses [53].…”