DEVELOPMENT AND EVALUATION OF ANTIGEN CAPTURE ELISA FOR THE DETECTION OF INFLUENZA VIRUS A

Mahshid Aria, Masoud Reza Seifiabad Shapouri, Mohammad Roayaei Ardakani, Hossein Motamedi, Azadeh Yektaserasht, Amin Jaidari

Abstract


The aim of the present study was to develop an Antigen Capture ELISA (AC-ELISA) for the diagnosis of avian influenza virus infections. For this purpose, the nucleoprotein (NP) of the virus was captured by a monoclonal antibody (D'C4) and then detected using a rabbit polyclonal antibody. The developed AC-ELISA did not show cross-reaction with other viral and bacterial pathogens of poultry, while it was able to detect H9 serotype of avian influenza virus as well as H1 and H3 types of human influenza viruses. The sensitivity of this AC-ELISA for the detection of an H9 avian influenza virus strain H9N2 (A/Chicken/Iran/AH-1/06) was 10 times greater than a hemagglutination assay and was comparable with the sensitivity of the RT-PCR method. Furthermore, this method could recognize the influenza virus in tracheal swabs of experimentally infected chickens following 3-5 days post-infection. Based on the obtained results, it can be concluded that the developed AC-ELISA is able to detect H9, H1, and H3 influenza virus serotypes and is sufficiently sensitive and specific for the detection of infections caused by H9 serotype but, its applicability, sensitivity, and specificity for the detection of other serotypes of the virus remain to be determined.

Key words: influenza virus A; nucleoprotein; monoclonal antibody; antigen capture ELISA

 

RAZVOJ IN PREIZKUS NOVE METODE ELISA Z UJETJEM PROTITELES ZA ODKRIVANJE VIRUSA INFLUENCE A

Namen raziskave je bil razvoj nove metode elisa z ujetjem protiteles za lažje diagnosticiranje okužbe z virusom ptičje influence. Nukleoprotein (NP) virusa smo najprej vezali z monoklonskimi protitelesi D’C4 in nato kompleks nukleoproteina in monoklonskega protitelesa označili s kunčjimi poliklonskimi protitelesi. Ta metoda ni kazala nobene navzkrižne (napačne) reakcije z drugimi virusi ali bakterijami ptičjega porekla, z njo pa smo lahko zaznali prisotnost virusa ptičje influence serotipa H9, pa tudi viruse človeške influence seroptipov H1 in H3. Občutljivost te metode za določanje linije H9N2 (A/Chicken/Iran/AH-1/106) virusa H9 je bila desetkrat višja v primerjavi z metodo hemaglutinacije in je bila primerljiva z občutljivostjo metode RT PCR. Z njo smo virus lahko določili v sapničnih izpirkih že 3 do 5 dni po okužbi poskusno okuženih piščancev. Iz pridobljenih rezultatov sklepamo, da nanovo razvita metoda elisa z ujetjem protiteles lahko zazna serotype H9, H1 in H3 virusa influence pri piščancih in ljudeh. Dokazali smo, da je metoda zanesljiva in zelo občutljiva za zaznavanje okužb s serotipom H9, uporabnost te nove metode za zaznavanje okužb z drugimi serotipi virusov influence pa bomo morali še dokazati.

Ključne besede: influenca virus A; nulleoprotein; monoklonska protitelesa; elisa z ujetjem protiteles


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DOI: http://dx.doi.org/10.26873/SVR-261-2017

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