As influenza viruses have developed resistance towards current drugs, it is urgent to find potential novel antiviral inhibitors. Here we generated an influenza virus reporter cell line in which the luciferase gene was driven by the influenza virus promoter and screened a small compound library (NCI Diversity Set II). Ten compounds were identified to have inhibitory activity against influenza A virus H1N1. Among them, four compounds blocked influenza virus replication through inhibiting the activity of vRNP. The compound NSC 335506 inhibited HA-mediated membrane fusion. It showed the inhibitory activity against H1N1, H9N2 and H5N1 subtype but not H3N2. Our results demonstrated that influenza virus reporter cell is a very useful tool to identify novel inhibitors against influenza A virus.