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目的评价甘露寡糖修饰布氏菌Rsα蛋白的生物活性。方法经1-乙基-3-(3-二甲基氨丙基)碳二亚胺(EDC)/N-羟基琥珀酰亚胺(NHS)化学偶联法,用4种甘露寡糖(甘露三糖、甘露四糖、甘露五糖及甘露六糖)对Rsα蛋白按不同摩尔比进行寡糖修饰。FITC荧光标记4种甘露寡糖修饰Rsα蛋白后,作用于RAW264.7细胞,荧光显微镜观察细胞对荧光的吞噬情况;流式细胞术检测细胞对蛋白的吞噬率。结果甘露寡糖与蛋白投料摩尔比达1 000∶1时获得的反应产物较稳定。甘露五糖修饰的Rsα蛋白被巨噬细胞吞噬的速度最快,其细胞吞噬率最高(25.89%)。结论筛选出五糖修饰的Rsα蛋白具有较高的生物活性,为研究其作为布氏菌糖蛋白候选疫苗的可行性奠定了基础。
Objective To evaluate the biological activity of Rsα protein of mannose oligosaccharide-modified Brucella. Methods Four kinds of mannooligosaccharides (mannose oligosaccharides, mannose, mannose, mannose, mannose, Trisaccharide, mannotetraose, mannopentose and mannosehexaose) were modified with oligosaccharides in different molar ratios to Rsα protein. FITC fluorescence-labeled 4 kinds of mannose oligosaccharides modified Rsα protein, the role of RAW264.7 cells, fluorescence microscope to observe the phagocytosis of cells; flow cytometry to detect the rate of protein phagocytosis. Results When the molar ratio of mannooligosaccharides to protein was 1 000:1, the reaction products obtained were more stable. Mannose pentose modified Rsα protein was phagocytosed by macrophages the fastest, the highest rate of cell phagocytosis (25.89%). Conclusion The pentasaccharide-modified Rsα protein was screened for its high biological activity, which laid the foundation for the study of its feasibility as a vaccine candidate for Brucella glycoprotein.