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    Development of a recombinant ELISA using yeast (Pichia pastoris)-expressed polypeptides for detection of antibodies against avian influenza A subtype H5 (2012)

    Art
    Zeitschriftenartikel / wissenschaftlicher Beitrag
    Autoren
    Shehata, A A
    Fiebig, P
    Sultan, H
    Hafez, H M (WE 15)
    Liebert, U G
    Quelle
    Journal of Virological Methods; 180(1/2) — S. 18–25
    ISSN: 0166-0934
    Sprache
    Englisch
    Verweise
    DOI: 10.1016/j.jviromet.2011.12.004
    Pubmed: 22197190
    Kontakt
    Institut für Geflügelkrankheiten

    Königsweg 63
    14163 Berlin
    Tel.+49 30 838 62676 Fax.+49 30 838 62690
    email:gefluegelkrankheiten@vetmed.fu-berlin

    Abstract / Zusammenfassung

    Two truncated sequences (designated P1 and rHA1) of influenza A virus subtype H5 haemagglutinin (HA) were cloned and expressed in yeast Pichia pastoris (P. pastoris). These polypeptides were used in an indirect recombinant ELISA (rELISA) for detection of H5 antibodies in poultry. Serum samples obtained from broiler chickens vaccinated with commercial inactivated vaccine (H5N2) and control negative sera from non-vaccinated chickens against influenza were tested using rP1-ELISA, rHA1-ELISA, whole H5N1-ELISA, Western blot, agar gel immunodiffusion (AGID) and haemagglutination inhibition (HI) tests. The rHA1-ELISA proved to be highly sensitive and specific. To study the validity of rHA1-ELISA, a total of 179 serum samples obtained from commercial broiler chickens vaccinated previously with commercial H5N2 inactivated vaccines, were tested by rHA1-ELISA, commercial ELISA (cELISA) and HI. The relative sensitivity and specificity between rHA1-ELISA, and HI tests were 100% and 70%, respectively, and between cELISA and HI were 100% and 57%, respectively. The agreement ratio between rHA1-ELISA and HI was 84.9% and between cELISA and HI tests was 76.5%. Serum samples obtained from ducks vaccinated with commercial inactivated H5N2 were tested by rHA1-ELISA and the results showed significant reactivity with duck sera. In conclusion, the results demonstrate the potential applicability of the rELISA for the determination of antibodies to H5 influenza virus in chickens and ducks.