Broad-spectrum detection of H5 subtype influenza A viruses with a new fluorescent immunochromatography system

Akira Sakurai; Katsuyoshi Takayama; Namiko Nomura; Tsubasa Munakata; Naoki Yamamoto; Tsuruki Tamura; Jitsuho Yamada; Masako Hashimoto; Kazuhiko Kuwahara; Yoshihiro Sakoda; Yoshihiko Suda; Yukuharu Kobayashi; Nobuo Sakaguchi; Hiroshi Kida; Michinori Kohara; Futoshi Shibasaki

doi:10.1371/journal.pone.0076753

Broad-spectrum detection of H5 subtype influenza A viruses with a new fluorescent immunochromatography system

PLoS One. 2013 Nov 6;8(11):e76753. doi: 10.1371/journal.pone.0076753. eCollection 2013.

Authors

Akira Sakurai¹, Katsuyoshi Takayama, Namiko Nomura, Tsubasa Munakata, Naoki Yamamoto, Tsuruki Tamura, Jitsuho Yamada, Masako Hashimoto, Kazuhiko Kuwahara, Yoshihiro Sakoda, Yoshihiko Suda, Yukuharu Kobayashi, Nobuo Sakaguchi, Hiroshi Kida, Michinori Kohara, Futoshi Shibasaki

Affiliation

¹ Department of Molecular Medical Research, Tokyo Metropolitan Institute of Medical Science, Kamikitazawa, Setagaya-ku, Tokyo, Japan.

Abstract

Immunochromatography (IC) is an antigen-detection assay that plays an important role in the rapid diagnosis of influenza virus because the protocol is short time and easy to use. Despite the usability of IC, the sensitivity is approximately 10(3) pfu per reaction. In addition, antigen-antibody interaction-based method cannot be used for the detection of influenza viruses with major antigenic change. In this study, we established the use of fluorescent immunochromatography (FLIC) to detect a broad spectrum of H5 subtype influenza A viruses. This method has improved sensitivity 10-100 fold higher than traditional IC because of the use of fluorescent conjugated beads. Our Type-E FLIC kit detected all of the H5 subtype influenza viruses that were examined, as well as recombinant hemagglutinin (HA) proteins (rHAs) belonging to the Eurasian H5 subtype viruses and the Type-N diagnosed North American H5 subtype influenza A viruses. Thus, this kit has the improved potential to detect H5 subtype influenza viruses of different clades with both Type-E and Type-N FLIC kits. Compared with PCR-based diagnosis, FLIC has a strong advantage in usability, because the sample preparation required for FLIC is only mix-and-drop without any additional steps such as RNA extraction. Our results can provide new strategies against the spread and transmission of HPAI H5N1 viruses in birds and mammals including humans.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Animals
Antibodies, Immobilized / chemistry
Antibodies, Viral / chemistry
Antibody Specificity
Chromatography, Affinity
Dogs
Hemagglutinin Glycoproteins, Influenza Virus / immunology
Humans
Immunoassay
Influenza A Virus, H5N1 Subtype / immunology
Influenza A Virus, H5N1 Subtype / isolation & purification*
Influenza, Human / diagnosis*
Influenza, Human / virology
Limit of Detection
Madin Darby Canine Kidney Cells
Reagent Kits, Diagnostic
Spectrometry, Fluorescence

Substances

Antibodies, Immobilized
Antibodies, Viral
Hemagglutinin Glycoproteins, Influenza Virus
Reagent Kits, Diagnostic
hemagglutinin, human influenza A virus

Grants and funding

This work was supported by grants from Ministry of Economy, Trade, and Industry, Japan as a part of “Program to support collaboration between hospitals and businesses for development and improvement of medical equipment and devices to solve unmet medical needs” (supplementary budget, 2010FY), Japan Society for the Promotion of Science KAKENHI (Grant Numbers 23659069 and 23659314), a Grant-in-Aid from the Ministry of Education, Culture, Sports, Science and Technology (MEXT), and a contract research fund from the Program of Founding Research Centers for Emerging and Reemerging Infectious Diseases (MEXT) to N.S. and partly by a grant for the Adaptable and Seamless Technology Transfer Program through Target-driven R&D (Japan Science and Technology Agency) to K.K. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.