Development of dual-function ELISA for effective antigen and antibody detection against H7 avian influenza virus

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.

Both Neutralizing and Non-Neutralizing Human H7N9 Influenza Vaccine-Induced Monoclonal Antibodies Confer Protection

“…The Gly137 residue was conserved in H7 strains from both lineages, explaining the broader binding pattern of 07-5G01 (Figure 1). This mutation has been previously described in the literature (He et al, 2013). The R65K substitution was identified in the escape mutants generated with 07-5F01.…”

Potency determination of inactivated H7 influenza vaccines using monoclonal antibody‐based ELISA and biolayer interferometry assays

“…The locations of these escape mutations, as well as those for 5A617 (previously mapped to antigenic site A – R131G), are shown in Figure 1. Escape mutant viruses with amino acid changes at positions 119 (G119E) and 157 (K157E) have previously been reported for mAbs 9822 and 62,21 respectively. In several attempts at isolating escape mutants to these two mAbs, viruses with both amino acid changes (G119E and K157E) were always obtained.…”

“…In order to broaden the H7 HA epitope representation of our mAbs, we generated and characterized additional panels of mAbs using approaches designed to select for mAbs directed to epitopes other than antigenic site A. In addition, we evaluated some existing mAbs (mAbs 62 and 98) developed to an older H7N1 strain to determine how well these mAbs would bind the more recent A(H7N9) hemagglutinins 21, 22. Monoclonal antibodies were assessed for binding to HA in an ELISA using inactivated A(H7N9) A/Shanghai/2/2013 virus.…”

Genotyping and detection of common avian and human origin-influenza viruses using a portable chemiluminescence imaging microarray

“…Some molecular diagnostic technologies have been used to diagnose and subtype influenza viruses: antigens and antibody detection (Duman et al 2013; He et al 2013), real-time PCR (Choi et al 2013; Dawood et al 2009; Gao et al 2013a; Hackett et al 2014; Poon et al 2009; Templeton et al 2004), sequencing (Deng et al 2011; Ghedin et al 2011; Rutvisuttinunt et al 2013), and microarray (Gall et al 2009; Han et al 2008; Heil et al 2010; Ryabinin et al 2011). In the assay described here, a chemiluminescence (CL) detection oligonucleotide microarray was developed and used to detect avian influenza A (H7N9), avian influenza A (H5N1), 2009 influenza A (H1N1), seasonal influenza A (H1N1), and seasonal influenza A (H3N2) by genotyping.…”