title: Development of a novel rapid immunochromatographic test specific for the H5 influenza virus.
abstract: Three anti-H5 influenza virus monoclonal antibody (mAb) clones, IFH5-26, IFH5-115 and IFH5-136, were obtained by immunising a BALB/C mouse with inactivated A/duck/Hokkaido/Vac-1/04 (H5N1). These mAbs were found to recognise specifically the haemagglutinin (HA) epitope of the influenza H5 subtypes by western blotting with recombinant HAs; however, these mAbs have no neutralising activity for A/duck/Hokkaido/84/02 (H5N3) or A/Puerto Ric/8/34 (H1N1). Each epitope of these mAbs was a conformational epitope that was formed from the regions located between 46 to 60 amino acids (aa) and 312 to 322 aa for IFH5-115, from 101 to 113 aa and 268 to 273 aa for IFH5-136 and from 61 to 80 aa and 290 to 300 aa for IFH5-26. The epitopes were located in the loop regions between the receptor region and alpha-helix structure in haemagglutinin 1 (HA1). Influenza A virus H5-specific rapid immunochromatographic test kits were tested as solid phase antibody/alkaline phosphate-conjugated mAb in the following three combinations: IFH5-26/IFH5-115, IFH5-136/IFH5-26 and IFH5-136/IFH5-115. In every combination, only influenza A H5 subtypes were detected. For effective clinical application, rapid dual discrimination immunochromatographic test kits in combination with H5 HA-specific mAb, IFA5-26 and IFA5-115 and the influenza A NP NP-specific mAb, FVA2-11, were developed. The dual discrimination immunochromatographic tests kits detected influenza A virus H5 subtypes as H5 line-positive and all influenza A subtypes as A line-positive simultaneously. The dual discrimination immunochromatographic test kits may be useful for discriminating highly pathogenic avian influenza A H5N1 viruses from seasonal influenza A virus, as well as for confirming influenza infection status in human, avian and mammalian hosts.
authors.affiliation: Ube Development Group, Fundamental Research Department, Fujirebio Inc., Setohara Danchi, Yoshiwa, Ube-shi 759-0134, Japan. ej-miyagaw@fujirebio.co.jp
authors.forename: Eiji
authors.initials: E
authors.lastname: Miyagawa
authors.forename: Hiroyuki
authors.initials: H
authors.lastname: Kogaki
authors.forename: Yoshiaki
authors.initials: Y
authors.lastname: Uchida
authors.forename: Nobuyuki
authors.initials: N
authors.lastname: Fujii
authors.forename: Takashi
authors.initials: T
authors.lastname: Shirakawa
authors.forename: Yoshihrio
authors.initials: Y
authors.lastname: Sakoda
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Kida
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000302:isolation & purification; D000914:Antibodies, Viral / Q000302:isolation & purification; D001717:Birds; D019267:Hemagglutinin Glycoproteins, Influenza Virus / Q000276:immunology / Q000302:isolation & purification*; D006801:Humans; D007118:Immunoassay / Q000379:methods; D005585:Influenza in Birds / Q000175:diagnosis* / Q000821:virology; D007251:Influenza, Human / Q000175:diagnosis* / Q000821:virology; D008322:Mammals; D051379:Mice; D008807:Mice, Inbred BALB C; D009976:Orthomyxoviridae Infections / Q000175:diagnosis* / Q000821:virology; D014773:Virology / Q000379:methods*
chemical_list: D000911:Antibodies, Monoclonal; D000914:Antibodies, Viral; D019267:Hemagglutinin Glycoproteins, Influenza Virus; C109963:hemagglutinin, avian influenza A virus