Ammonium acetate, ammonium hydroxide, N-acetyl-D-neuraminic acid (C11H19NO9, MW 309.3 Da), α-2,3-sialyllactose, α-2,6-sialyllactose, and fetuin from fetal calf serum were obtained from Sigma-Aldrich (St. Louis, MO). N-Acetyl-D-[1,2,3-13C3]-neuraminic acid internal standard (13C3C8H19NO9, MW 312.3 Da) was acquired from Omicron Biochemicals, Inc. (South Bend, IN). All chemicals were of the highest purity available and used without further purification. NA-Fluor 2× Assay buffer, 66.6 mM 2-(N-morpholino) ethanesulfonic acid (MES) buffer, 8 mM CaCl2, pH 6.5, was from Applied Biosystems (Foster City, CA). The LC column was from Imtakt USA (Portland, OR), and influenza vaccines A–F (Table 1 ) were commercially available.
During the implementation of the fluorescence method, the NA-Fluor Influenza Neuraminidase Assay kit from Applied Biosystems, Cat No. 4457091, was used. Ethanol absolute, 4-methylumbelliferone (4-MU), and sodium chloride solution were from Sigma-Aldrich. 96-well opaque black flat-bottom plates from Corning (Corning, NY) were used with a SpectraMax M5e micro plate reader from Molecular Devices LLC (Sunnyvale, CA) for fluorescence measurements.
During the implementation of the fluorescence method, the NA-Fluor Influenza Neuraminidase Assay kit from Applied Biosystems, Cat No. 4457091, was used. Ethanol absolute, 4-methylumbelliferone (4-MU), and sodium chloride solution were from Sigma-Aldrich. 96-well opaque black flat-bottom plates from Corning (Corning, NY) were used with a SpectraMax M5e micro plate reader from Molecular Devices LLC (Sunnyvale, CA) for fluorescence measurements.