Abstract
Influenza A and Sendai viruses bind toneolacto-series gangliosides isolated from human granulocytes. Differences in receptor specificity of influenza viruses A/PR/8/34 (H1N1), A/X-31 (H3N2), and parainfluenza Sendai virus (HNF1, Z-strain) were determined by two direct solid phase binding assays: the overlay technique, which combines high-resolution in the separation of gangliosides on thin-layer chromatograms with direct binding; and the microwell adsorption assay as a convenient binding assay which is performed in microtitre wells to estimate the avidity of binding to an isolated ganglioside. Both methods were applied for comparative binding studies. Viruses were found to exhibit specificity for oligosaccharides and sialic acids as well as for chain length of the neutral carbohydrate backbone, whereas differing fatty acids (C24:1 and C16:0) in the ceramide portion had no impact on virus adsorption. Terminal sialyloligosaccharides Neu5Acα2-3Galβ1-4Glc-R of GM3, and Neu5Acα2-3Galβ1-4GlcNAc-R as well as Neu5Acα2-6Galβ1-4GlcNAc-R ofneolacto-series gangliosides with nLcOse4Cer and nLcOse6Cer backbone, exhibited significant specific receptor activity towards the different viruses. To compare the data revealed from both test systems, values of virus binding were ascertained by a non-parametric statistical approach based on rank correlation. The rank correlation coefficientr s was calculated according to Spearman from each virus binding towards GM3, IV3Neu5Ac-nLcOse4Cer, IV6Neu5Ac-nLcOse4Cer and VI3Neu5Ac-nLcOse6SCer. The rank correlation coefficients 0.74, 0.95 and 0.92, which were determined for A/PR/8/34 (H1N1), A/X-31 (H3N2) and Sendai virus (HNF1, Z-strain), respectively, indicated that both assays generate highly correlated experimental data. Based on these results, analyses of virus binding on thin-layer chromatograms as well as in microwells were found equivalent tools for ganglioside receptor studies.
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Abbreviations: BSA, bovine serum albumin; GSL(s), glycosphingolipids; HPTLC, high performance thin-layer chromatography; PBS, phosphate buffered saline; Neu5Ac,N-acetylneuraminic acid [35];r s = rank correlation coefficient according to Spearman. The designation of the glycosphingolipids follows the IUPAC-IUB recommendations [36]. LacCer or lactosylceramide, Galβ1-4Glcβ1-1Cer; lacto-N-neotetraosylceramide or nLcOse4Cer, Galβ1-4GlcNAcβ1-3Galβ1-4Glcβ1-1Cer; lacto-N-norhexaosylceramide or nLcOse6Cer, Galβ1-4GlcNAcβ1-3Galβ1-4GlcNAcβ1-3Galβ1-4Glcβ1-1Cer; GM3 (according to Svennerholm [37]) or II3Neu5AcLacCer.
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Müthing, J., Unland, F. A comparative assessment of TLC overlay technique and microwell adsorption assay in the examination of influenza A and Sendai virus specificities towards oligosaccharides and sialic acid linkages of gangliosides. Glycoconjugate J 11, 486–492 (1994). https://doi.org/10.1007/BF00731285
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DOI: https://doi.org/10.1007/BF00731285