Multispecies detection of antibodies to influenza A viruses by a double-antigen sandwich ELISA.
A double-antigen sandwich ELISA was developed for the detection of antibodies to influenza A viruses. A recombinant nucleoprotein (rNP) of influenza A virus was used as a capture antigen and an HRP-conjugate for detecting the antibodies. A total of 125 serum samples from birds of different species including chickens, geese, open-billed storks, Khaki Campbell ducks, lesser whistling ducks, and pigeons with known antibodies were tested by ELISA. The sensitivity and the specificity of ELISA were found to be 98% and 97.3%, respectively. The assay was able to detect the presence of influenza A antibodies as early as the fourth day post-inoculation in ducks infected experimentally with influenza A (H5N1) virus. Excellent agreement (97.6%) was obtained between this sandwich ELISA and the hemagglutination inhibition (HI) tests (kappa=0.95). The double-antigen sandwich ELISA correlated well with a commercial avian influenza (AI) multispecies ELISA and was slightly more sensitive than the AI multispecies ELISA. These findings indicate that the double-antigen sandwich ELISA based on rNP may offer an effective screening method for serodiagnosis of influenza A virus. The double-antigen sandwich ELISA also enables the detection of antibodies to influenza A viruses in different species without the need for species-specific secondary antibodies.
Watcharatanyatip
Kamolwan
K
Department of Immunology, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok 10700, Thailand.
Boonmoh
Sirikwan
S
Chaichoun
Kridsada
K
Songserm
Taweesak
T
Woratanti
Mingkhwan
M
Dharakul
Tararaj
T
D000818:Animals; D000914:Antibodies, Viral / Q000097:blood*; D000956:Antigens, Viral / Q000235:genetics; D001715:Bird Diseases / Q000175:diagnosis*; D001717:Birds; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D006385:Hemagglutination Inhibition Tests; D009980:Influenza A virus / Q000276:immunology*; D005585:Influenza in Birds / Q000175:diagnosis*; D019590:Nucleocapsid Proteins; D011200:Poultry; D011201:Poultry Diseases / Q000175:diagnosis*; D016601:RNA-Binding Proteins / Q000235:genetics; D011994:Recombinant Proteins / Q000235:genetics; D012680:Sensitivity and Specificity; D013223:Statistics as Topic; D014758:Viral Core Proteins / Q000235:genetics
D000914:Antibodies, Viral; D000956:Antigens, Viral; C048842:NP protein, Influenza A virus; D019590:Nucleocapsid Proteins; D016601:RNA-Binding Proteins; D011994:Recombinant Proteins; D014758:Viral Core Proteins