Open Access Open Badges Research article

Development of a sensitive novel diagnostic kit for the highly pathogenic avian influenza A (H5N1) virus

Yasuko Tsunetsugu-Yokota12*, Kengo Nishimura3, Shuhei Misawa3, Mie Kobayashi-Ishihara1, Hitoshi Takahashi4, Ikuyo Takayama4, Kazuo Ohnishi1, Shigeyuki Itamura4, Hang LK Nguyen5, Mai TQ Le5, Giang T Dang6, Long T Nguyen6, Masato Tashiro4 and Tsutomu Kageyama4

Author Affiliations

1 Department of Immunology, National Institute of Infectious Diseases, Shinjuku Tokyo 162-8640, Japan

2 Department of Medical Technology, School of Human Sciences, Tokyo University of Technology, 5-23-22 Nishi-Kamata, Ohta-ku Tokyo 144-8535, Japan

3 Tsuruga Institute of Biotechnology, Toyobo, Co., Ltd., Fukui 914-8550, Japan

4 National Institute of Infectious Diseases, Influenza Virus Research Center, Musashimurayama Tokyo 208-0011, Japan

5 Department of Virology, National Institute of Hygiene and Epidemiology, Hanoi, Vietnam

6 Pasteur Institute, National Influenza Center, Ho Chi Minh City, Vietnam

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BMC Infectious Diseases 2014, 14:362  doi:10.1186/1471-2334-14-362

Published: 3 July 2014



Sporadic emergence of the highly pathogenic avian influenza (HPAI) H5N1 virus infection in humans is a serious concern because of the potential for a pandemic. Conventional or quantitative RT-PCR is the standard laboratory test to detect viral influenza infections. However, this technology requires well-equipped laboratories and highly trained personnel. A rapid, sensitive, and specific alternative screening method is needed.


By a luminescence-linked enzyme immunoassay, we have developed a H5N1 HPAI virus detection kit using anti-H5 hemagglutinin monoclonal antibodies in combination with the detection of a universal NP antigen of the type A influenza virus. The process takes 15 minutes by use of the fully automated luminescence analyzer, POCube.


We tested this H5/A kit using 19 clinical specimens from 13 patients stored in Vietnam who were infected with clade 1.1 or clade 2.3.4 H5N1 HPAI virus. Approximately 80% of clinical specimens were H5-positive using the POCube system, whereas only 10% of the H5-positive samples were detected as influenza A-positive by an immunochromatography-based rapid diagnostic kit.


This novel H5/A kit using POCube is served as a rapid and sensitive screening test for H5N1 HPAI virus infection in humans.

H5 hemagglutinin; Highly pathogenic avian influenza virus; Rapid influenza diagnosis; Monoclonal antibody; Clinical specimens