Enzyme-linked immunosorbent assays (ELISAs) were performed to test the cross-reactivity of the serum samples against multiple influenza A subtypes. Recombinant HA proteins, including H1 (A/New York/18/2009(H1N1), H1-NY18), H3 (A/ Aichi2/68 (H3N2), H3-A2/68), H5 (A/Vietnam/1203/04 (H5N1), H5-VN04), and H7 (A/Netherlands 219/03 (H7N7), H7-NL03), were expressed in insect cells as trimers of HA ectodomains [1] . HA proteins (0.2 lg) were coated onto 96-well Maxisorb ELISA plate (Nunc) at 2 lg/mL in phosphate-buffered saline (PBS) at 4°C overnight. The plate was washed with PBS for 3 times to remove uncoated proteins. Serially diluted serum samples were applied to the HA-coated plates, followed by Horseradish Peroxidase (HRP)-anti-human IgG or IgM (Pierce Biotechnology), to detect the IgGs or IgMs against various HA subtypes in the serum samples. The optical density at 450 nm was measured after incubation of the peroxidase tetramethylbenzidine substrate system.