Development of mouse monoclonal antibody for detecting hemagglutinin of avian influenza A(H7N9) virus and preventing virus infection

Abstract

Many cases of avian influenza A(H7N9) virus infection in humans have been reported since its first emergence in 2013. The disease is of concern because most patients have become severely ill with roughly 30% mortality rate. Because the threat in public health caused by H7N9 virus remains high, advance preparedness is essentially needed. In this study, the recombinant H7N9 hemagglutinin (HA) was expressed in insect cells and purified for generation of two monoclonal antibodies, named F3-2 and 1C6B. F3-2 can only recognize the H7N9 HA without having cross-reactivity with HA proteins of H1N1, H3N2, H5N1, and H7N7. 1C6B has the similar specificity with F3-2, but 1C6B can also bind to H7N7 HA. The binding epitope of F3-2 is mainly located in the region of H7N9 HA(299-307). The binding epitope of 1C6B is located in the region of H7N9 HA(489-506). F3-2 and 1C6B could not effectively inhibit the hemagglutination activity of H7N9 HA. However, F3-2 can prevent H7N9 HA from trypsin cleavage and can bind to H7N9 HA which has undergone pH-induced conformational change. F3-2 also has the ability of binding to H7N9 viral particles and inhibiting H7N9 virus infection to MDCK cells with the IC50 value of 22.18 μg/mL. In addition, F3-2 and 1C6B were utilized for comprising a lateral flow immunochromatographic test strip for specific detection of H7N9 HA. Key points• Two mouse monoclonal antibodies, F3-2 and 1C6B, were generated for recognizing the novel binding epitopes in H7N9 HA.• F3-2 can prevent H7N9 HA from trypsin cleavage and inhibit H7N9 virus infection to MDCK cells.• F3-2 and 1C6B were developed as a lateral flow immunochromatographic test for specific detection of H7N9 HA.

Neutralizing Monoclonal Antibodies Inhibit SARS-CoV-2 Infection through Blocking Membrane Fusion

“…The animal experiments were approved by the Institutional Animal Care and Use Committee of National Taiwan University (approval number: NTU-109-EL-00051) and implemented in accordance with the animal care and ethics guidelines. The procedures for generation of hybridomas were performed as described previously ( 53 , 54 ) with slight modifications. In brief, two BALB/c male mice were immunized one time with a mixture of S2 (100 μg) and complete Freund’s adjuvant (0.25 mL) and two times with a mixture of S2 (50 μg) and incomplete Freund’s adjuvant (0.25 mL) at a 2-week interval through intraperitoneal injection, followed by a final booster injection of S2 (50 μg) in 0.25 mL of PBS.…”

Neutralization or enhancement of SARS-CoV-2 infection by a monoclonal antibody targeting a specific epitope in the spike receptor-binding domain

“…The animal experiment was approved by the Institutional Animal Care and Use Committee (IACUC) of National Taiwan University (Approval Number: NTU-109-EL-00051) and implemented in accordance with the animal care and ethics guidelines. The procedures for hybridomas production were performed as described previously ( Cheng and Chang, 2021 ; Chiang et al, 2021 ) with slight modifications. In brief, two BALB/c male mice were immunized in a 2-week interval through intraperitoneal injection with a mixture of purified RBD (100 μg) and complete or incomplete Freund's adjuvant (0.25 mL) (Sigma-Aldrich), followed by a final booster injection of purified RBD (50 μg) in PBS.…”

Cross-reactive single-domain antibodies to hemagglutinin stem region protect mice from group 1 influenza A virus infection

“…The procedure was carried out as described previously, with minor modifications [49]. 1 µg/ml rFL HA in carbonate-bicarbonate buffer pH 9.6 was immobilized on the surface of 96-well polystyrene microplates (Corning Inc., NY, USA) overnight at 4°C.…”