DAS181 is a novel therapeutic candidate against influenza virus which functions via the mechanism of removing the virus receptor, sialic acid (Sia) from the adjacent glycan structures. DAS181 and its analogues have previously been shown to be potently active against multiple strains of seasonal and avian influenza virus strains in several experimental models including cell lines, mice and ferrets. Here we demonstrate that DAS181 treatment leads to desialylation of both the alpha2-6-linked and alpha2-3-linked Sia in the ex vivo human lung tissue culture and primary pneumocytes. DAS181 treatment also effectively protects the human lung tissue and pneumocytes against the highly pathogenic avian influenza (HPAI) H5N1 (A/Vietnam/3046/2004). Two doses of DAS181 treatment given at 12 h apart were sufficient to block H5N1 infection in the ex vivo lung tissue culture. These findings support the potential value of DAS181 as a broad-spectrum therapeutic agent against influenza viruses, especially H5N1.