Since March 2013, the H7N9 subtype of avian influenza virus (AIV) has become an important zoonotic infectious disease, garnering significant global attention because of its potential to affect human health. Establishing a rapid, effective, and sensitive method to detect H7 subtype AIVs is crucial for disease control. In this study, we developed a graphene oxide multilayer quantum dot-based immunochromatographic strip for the ultrasensitive detection of H7 subtype AIVs. The method demonstrated excellent sensitivity, with a limit of detection of 0.063 hemagglutinin units and 0.016 ng/ml for the hemagglutinin protein. The method exhibited remarkable specificity, with no reaction with other subtypes of influenza A virus andno cross-reactivity with other types of avian virus. Additionally, this method exhibited excellent reproducibility, with both inter-group and intra-group variations remaining below 10 %. Preliminary testing on avian clinical samples showed impressive consistency, underscoring the method's reliability. These initial results suggest that this detection approach has significant potential for widespread use in analyzing avian clinical samples, indicating substantial promise for its future application in various diagnostic settings.