The performance of a simplified nucleoprotein (NP) and hemagglutinin- subtype-9 (H9) based reverse transcriptase-polymerase chain reaction-enzyme linked immunosorbent assay (RT-PCR-ELISA) for the detection of avian influenza virus (AIV) subtype H9N2 was compared to the standard the virus isolation method and serology testing using hemagglutination (HA) and hemagglutination inhibition (HI) tests. The H9-based RT-PCR-ELISA was 100% sensitive when compared to virus isolation method in detecting H9N2 from experimentally infected specific-pathogen-free (SPF) chickens. The NP- and H9-based RT-PCR-ELISA have a detection limit similar to the virus isolation method in detecting serially diluted tracheal swab samples obtained from chickens inoculated with H9N2. Both RT-PCR-ELISAs were also ten times more sensitive than agarose gel electrophoresis for the detection of PCR products. The result of this study demonstrate that the developed RT-PCR-ELISA is a simple and sensitive assay for the detection of type A influenza virus, particularly AIV subtype H9N2, in chickens.