Abstract
BackgroundOutbreaks in poultry involving influenza virus from H7 subtype have resulted in human infections, thus causing a major concern for public health, as well as for the poultry industry. Currently, no efficient rapid test is available for large-scale detection of either antigen or antibody of H7 avian influenza viruses.ResultsIn the present study, a dual function ELISA was developed for the effective detection of antigen and antibody against H7 AIVs. The test was established based on antigen-capture-ELISA and epitope blocking ELISA. The two Mabs 62 and 98 which were exploited in the assay were identified to recognize two conformational neutralizing epitopes on H7 HA1. Both of the epitopes exist in all of the human H7 strains, including the recent H7N9 strain from China and > 96.6% of avian H7 strains. The dual ELISA was able to detect all of the five H7 antigens tested without any cross reaction to other influenza subtypes. The antigen detection limit was less than 1 HA unit of H7. For antibody detection, the sensitivity and specificity of the dual ELISA was evaluated and compared to HI and microneutralization using immunized animal sera to different H7 strains and different subtypes of AIVs. Results indicated that antibodies to H7 were readily detected in immunized animal sera by the dual ELISA whereas specimens with antibodies to other AIVs yielded negative results.ConclusionsThis is the first dual-function ELISA reported for either antigen or antibody detection against H7 AIVs. The assay was highly sensitive and 100% specific in both functions rendering it effective for H7 diagnosis.
Highlights
Outbreaks in poultry involving influenza virus from H7 subtype have resulted in human infections, causing a major concern for public health, as well as for the poultry industry
The results indicated that for the first time, antigen and antibody detection could be performed with the same device and Mabs for specific and sensitive H7 AIV detection
All animal sera from H7 immunization, including chicken, mouse and guinea pigs, showed positive blocking in the dual ELISA, indicating the assay is effective for sera from any species. These results indicate that the antibody detection in the dual ELISA could positively identify serum samples containing antibodies to H7 without any cross reaction to sera from other subtypes
Summary
Outbreaks in poultry involving influenza virus from H7 subtype have resulted in human infections, causing a major concern for public health, as well as for the poultry industry. Conventional laboratory methods for influenza virus detection include virus isolation in embryonated eggs or Madin-Darby canine kidney (MDCK) cells, followed by subsequent HA subtype identification using serological methods. Molecular detection methods such as real-time PCR assays have been widely applied for the laboratory diagnosis of influenza infections [6,7] and HA subtype identification [8]. Both conventional and laboratory methods are technically demanding and are not suitable for on-site use in field investigations. The development of rapid H7 subtype influenza virus detection tests in dot ELISA (enzyme-linked immunosorbent assay) [9], AC-ELISA (antigen-capture ELISA), and chromatographic strip formats [10] using H7 monoclonal antibodies (MAbs) is preferred
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