Combining magnetic nanoparticle with biotinylated nanobodies for rapid and sensitive detection of influenza H3N2
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NANO EXPRESS
Open Access
Combining magnetic nanoparticle with biotinylated nanobodies for rapid and sensitive detection of influenza H3N2 Min Zhu1,2, Yonghong Hu3, Guirong Li2, Weijun Ou2, Panyong Mao4, Shaojie Xin4* and Yakun Wan1,2,5*
Abstract Our objective is to develop a rapid and sensitive assay based on magnetic beads to detect the concentration of influenza H3N2. The possibility of using variable domain heavy-chain antibodies (nanobody) as diagnostic tools for influenza H3N2 was investigated. A healthy camel was immunized with inactivated influenza H3N2. A nanobody library of 8 × 108 clones was constructed and phage displayed. After three successive biopanning steps, H3N2-specific nanobodies were successfully isolated, expressed in Escherichia coli, and purified. Sequence analysis of the nanobodies revealed that we possessed four classes of nanobodies against H3N2. Two nanobodies were further used to prepare our rapid diagnostic kit. Biotinylated nanobody was effectively immobilized onto the surface of streptavidin magnetic beads. The modified magnetic beads with nanobody capture specifically influenza H3N2 and can still be recognized by nanobodies conjugated to horseradish peroxidase (HRP) conjugates. Under optimized conditions, the present immunoassay exhibited a relatively high sensitive detection with a limit of 50 ng/mL. In conclusion, by combining magnetic beads with specific nanobodies, this assay provides a promising influenza detection assay to develop a potential rapid, sensitive, and low-cost diagnostic tool to screen for influenza infections. Keywords: Influenza A grade 2 (H3N2); Nanobody; Magnetic nanoparticles; Biotinylation
Background Influenza A and B viruses cause a pandemic threat to human health throughout the world [1]. Sporadic transmission of influenza viruses from birds to humans could lead to unpredictable pandemic outbreaks. Influenza is an infectious disease of the respiratory tract that can infect millions of people and kills hundreds of thousands of them [2]. Humans, infected with influenza A, manifest typically an acute upper respiratory tract illness characterized by fever, cough, and sore throat. Disease severity depends mainly on the virulence of the influenza virus strain and immune competence of the patients [3]. Influenza viruses are members of the Orthomyxoviridae family, and they are further classed as A, B, and C viruses [4]. Until now, 17 influenza A hemagglutinin (HA) subtypes have been described. However, only a limited * Correspondence: [email protected]; [email protected] 4 302 Military Hospital of China, Beijing 100039, People’s Republic of China 1 The Key Laboratory of Developmental Genes and Human Disease, Ministry of Education, Institute of Life Sciences, Southeast University, Nanjing 210096, People’s Republic of China Full list of author information is available at the end of the article
number of influenza A viruses (IAV), such as H1, H2, H3, H5, H6, H7, and H9, have been implicated with human infection [5]. The high mutational rate of the vir
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