A human strain of influenza virus (A, H1N1) was shown to bind in an unexpected way to leukocyte and other gangliosides when compared with avian virus (A, H4N6) as assayed on TLC plates. The human strain bound only to species with about 10 or more sugars, while the avian strain bound to a wide range of gangliosides including the 5-sugar gangliosides. By use of specific lectins, antibodies, and FAB and MALDI-TOF mass spectrometry an attempt was done to preliminary identify the sequences of leukocyte gangliosides recognized by the human strain. The virus binding pattern did not follow binding by VIM-2 monoclonal antibody and was not identical with binding by anti-sialyl Lewis x antibody. There was no binding by the virus of linear NeuAcalpha3- or NeuAcalpha6-containing gangliosides with up to seven monosaccharides per mol of ceramide. Active species were minor NeuAcalpha6-containing molecules with probably repeated HexHexNAc units and fucose branches. This investigation demonstrates marked distinctions in the recognition of gangliosides between avian and human influenza viruses. Our data emphasize the importance of structural factors associated with more distant parts of the binding epitope and the complexity of carbohydrate recognition by human influenza viruses.