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. 2015 Aug 12;5:13038. doi: 10.1038/srep13038

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Copyright © 2015, Macmillan Publishers Limited

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(a) pCG-HN and its mutants were constructed and transfected into BHK-21F cells. Total HN protein expression was verified by western blotting. (b) The amount of each HN protein at the cell surface (the cell surface expression; CSE) was determined by FACS analysis at 16 h post-transfection of BHK-21F cells by using anti-G7 serum. (c) The hemadsorption (HAd) activity was determined based on the ability of the HN expressed at the cell surface to adsorb chicken erythrocytes at 4 °C. (d) The neuraminidase (NA) activity was determined as the ability of the cell surface HA proteins to catalyze the release of sialic acid from 2’-(4-methylumbelliferyl)-α-D-N-acetylneuraminic acid. For all three of these assays, the background level obtained with vector alone was subtracted. All data are expressed relative to the amount of wt protein; bars represent the mean ± SD of three independent experiments (n = 3) (*P < 0.05, ***P < 0.001).