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首页> 外文期刊>Mycologia >FITC-lectin avidity of Cryptococcus neoformans cell wall and capsular components
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FITC-lectin avidity of Cryptococcus neoformans cell wall and capsular components

机译：新型隐球菌细胞壁和荚膜成分的FITC-凝集素亲和力

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Flow cytometry and confocal microscopy were used to quantify and visualize FITC-lectin binding to cell-surface carbohydrate ligands of log and stationary phase acapsular and capsular Cryptococcus neoformans strains. Cell populations demonstrated marked avidity for terminal alpha-linked mannose and glucose specific FITC-Con A, mannose specific FITC-GNL. as well as N-acetylglucosamine specific FITC-WGA. Exposure to other FITC-lectins specific for mannose, fucose and N-acetylgalactosamine resulted in little cell-surface fluorescence. The nature of cell-surface carbohydrates was investigated further by measurement of the fluorescence from surfaces of log and stationary phase cell populations after exposing them to increasing concentrations of FITC-Con A and FITC-WGA. Cell fluorescence increased significantly with small increases in FITC-Con A and FITC-WGA concentrations attaining reproducible maxima. Measurements of this nature supported calculation of the lectin binding determinants EC 50, Hn, Fmax and relative Bmax values. EC50 values indicated that the yeast-cell surfaces had greatest affinity for FITC-WGA, however, relative Bmax values indicated that greater numbers of Con A binding sites were present on these same cell surfaces. Hn values suggested a co-operative lectin-carbohydrate ligand interaction. Imaging of FITC-Con A and FITC-WGA cell-surface fluorescence by confocal microscopy demonstrated marked localization of both lectins to cell surfaces associated with cell division and maturation, indicative of dynamic carbohydrate ligand exposure and masking. Some fluorescence was associated with entrapment of FITC-Con A by capsular components, but FITC-Con A and FITC-WGA readily penetrated the capsule matrix to bind to the same cell surfaces labelled in acapsular cells.
机译：流式细胞术和共聚焦显微镜用于量化和可视化FITC-凝集素与log和固定相荚膜和荚膜隐球菌菌株的细胞表面碳水化合物配体的结合。细胞群显示出对末端α-连接的甘露糖和葡萄糖特异性FITC-Con A，甘露糖特异性FITC-GNL具有明显的亲和力。以及N-乙酰氨基葡萄糖专用FITC-WGA。暴露于对甘露糖，岩藻糖和N-乙酰半乳糖胺有特异性的其他FITC-凝集素导致细胞表面荧光很少。在暴露于浓度不断增加的FITC-Con A和FITC-WGA之后，通过测量对数和固定相细胞群体表面的荧光来进一步研究细胞表面碳水化合物的性质。随着FITC-Con A和FITC-WGA浓度的少量增加，细胞荧光显着增加，达到可重复的最大值。这种性质的测量结果支持计算凝集素结合决定簇EC 50，Hn，Fmax和相对Bmax值。 EC50值表明酵母细胞表面对FITC-WGA具有最大亲和力，但是，相对Bmax值表明在这些相同的细胞表面上存在大量的Con A结合位点。 Hn值表明了凝集素-碳水化合物配体的相互作用。通过共聚焦显微镜对FITC-Con A和FITC-WGA细胞表面的荧光成像显示，两种凝集素均明显定位于与细胞分裂和成熟相关的细胞表面，表明存在动态碳水化合物配体暴露和掩盖现象。某些荧光与荚膜成分截留FITC-Con A有关，但是FITC-Con A和FITC-WGA容易渗透到胶囊基质中，从而与荚膜细胞中标记的相同细胞表面结合。

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