Abstract
BackgroundH7N9 avian influenza virus (AIV) including highly and low pathogenic viruses have been detected in China since 2013. H7N9 AIV has a high mortality rate after infection in humans, and most human cases have close contacted with poultry in the live poultry market. Therefore, it is necessary to develop a rapid point-of-care testing (POCT) technique for H7N9 AIV detection.MethodsThe H7N9 AIV was inactivated and purified, and was used as the antigen to immunize BALB/c. Twelve H7-HA specific monoclonal antibodies (McAbs) were produced through the hybridoma technique. The McAb 10A8 was conjugated with colloid gold as detecting antibody; McAb 9B6 was dispensed on the nitrocellulose membran as the capture test line and the Goat-anti mouse IgG antibody was dispensed as control line respectively. The immunochromatographic strip was prepared.ResultsThe analysis of ELISA and virus neutralization test showed that the obtained McAbs specifically recognized H7 HA. Based on the prepared strip, the detection of H7 AIV was achieved within 10 min. No cross-reaction occurred between H7 AIVs and other tested viruses. The detection limit of the strip for H7 was 2.4 log10EID50/0.1 mL for chicken swab samples.ConclusionThe McAbs were specific for H7 and the immunochromatographic strip developed in this study was convenient, rapid and reliable for the detection of H7 AIV. The strip could provide an effective method for the rapid and early detection of H7 AIV.
Highlights
Human infected with H7N9 avian influenza virus (AIV) was first reported in the spring of 2013 in China [1, 2]
Preparation and characterization of monoclonal antibodies Twelve McAbs against H7N9 AIVs HA protein were produced, and only two McAbs 9B6 and 10A8 related to the strip were introduced in this article (Fig. 1)
The epitopes recognized by McAbs 9B6 and 10A8 were determined as GVTSACRRSGSSFYAEMK, and YKSTQSAIDQITGKLNRL, respectively (Table 3)
Summary
Human infected with H7N9 avian influenza virus (AIV) was first reported in the spring of 2013 in China [1, 2]. The National Avian Influenza Reference Laboratory (NAIRL) has established serological diagnostic techniques including hemagglutination (HA) and hemagglutination inhibition (HI) assays, agar gel immunodiffusion (AGID) assays, neuraminidase inhibition (NI) assays and indirect enzyme-linked immunosorbent assays (ELISA). Molecular diagnostic techniques include reverse transcriptionpolymerase chain reaction (RT-PCR) and real-time RT-PCR [13, 14]. These traditional detection methods are time-consuming, laborious with complicated operations, and prone to false positive results. H7N9 avian influenza virus (AIV) including highly and low pathogenic viruses have been detected in China since 2013. It is necessary to develop a rapid point-of-care testing (POCT) technique for H7N9 AIV detection
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