Avian influenza virus (AIV) as a type of highly pathogenic influenza A virus, can not only cause serious illness and death in poultry but also threat human health and lead to public panic. Rapid, sensitive detection of AIV is urgent and significant for prevention and timely control of influenza epidemics. Herein, we reported an inductively coupled plasma mass spectrometry (ICP-MS) based method for the analysis of AIV virions on the basis of the selective recognition between lectin Con A and glycoproteins on AIV surface and signal amplification of hybridization chain reaction (HCR). With H9N2 as the model AIV, the limit of detection was down to 0.12 ng mL−1 due to the dual amplification effect of AuNPs and HCR, and the linear range was 0.4–50 ng mL−1 with the relative standard deviation of 7.9% for seven replicate detections of 2 ng mL−1 H9N2 virions. Furthermore, the applicability of the method for the analysis of real biological samples was demonstrated by the spiking tests. The proposed approach is highly specific and sensitive for the detection of AIV with good application potential in early diagnosis, which is helpful for the prevention of influenza outbreak.