FRACTIONATION & CHARACTERIZATION OF OCULAR MUCUS

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• 批准号: 3259589
• 负责人: CHIEN-CHYOU CHAO
• 金额: $ 5.14万
• 依托单位: EYE AND EAR HOSPITAL OF PITTSBURGH
• 依托单位国家: 美国
• 财政年份: 1983
• 资助国家: 美国
• 起止时间: 1983-07-01 至 1987-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3259589
• 关键词: affinity chromatography; aminoacid analyzer; body fluid viscosity; carbohydrates; conjunctivitis; density gradient ultracentrifugation; electrofocusing; enzymes; gas chromatography; human tissue; immunoelectrophoresis; immunologic assay /test; ion exchange chromatography; keratoconjunctivitis; lipids; macromolecule; mass spectrometry; membrane activity; molecular weight; monoclonal antibody; mucins; mucopolysaccharides; mucus; nonblood rheology; nuclear magnetic resonance spectroscopy; proteoglycan; solubility; tear; thin layer chromatography; zonal electrophoresis

The object of the proposed research is to acquire information leading to a clearer understanding of the roles of various ocular mucus and tear components in mucus gel formation and tear film stability in health and disease. This will involve isolation and characterization of the responsible components in mucus from both normal and diseased eyes. Special attention will be paid to the isolation of ocular mucin. Other mucus/tear components to be studied include serum and tear specific proteins, lipids, glycosaminoglycans, and proteoglycans. For the purification of ocular mucin, specimens will be separated according to blood type of individual ocular mucus donors. Methods of isolation and purification will include selective solvent extraction, affinity and ion exchange chromatographies, gradient ultracentrifugation, and immunological methods. Chemical compositional analyses of isolates will include carbohydrates by GLC, amino acids and amino sugars by amino acid analysis, and lipid components by TLC and GLC. Structural analyses of mucin will involve amino acid sequencing of the peptide core, and GLC, MS, and NMR spectroscopy in conjunction with sugar analysis and immunological assays of the oligosaccharide chains. (Monoclonal) antibodies to mucin, tear prealbumin, and possibly de-glycosylated mucin isolates will be prepared. Physical property studies of individual mucus components, particularly mucin, will include measurements/determinations of viscosity, solubility, molecular weight, surface activity, and mode of binding among structural elements. Recent advances in microanalyses and our experience with the application of these techniques will provide an excellent opportunity for achieving these objectives.

拟议研究的目的是获取信息 从而更清楚地了解各种眼部的作用 粘液凝胶形成和泪膜中的粘液和泪液成分 健康和疾病的稳定性。 这将涉及隔离和 粘液中负责成分的表征 正常和患病的眼睛。 将特别关注 眼粘蛋白的分离。 其他粘液/泪液成分 所研究的包括血清和泪液特异性蛋白质、脂质、 糖胺聚糖和蛋白聚糖。 用于纯化 眼粘蛋白，标本将根据血型分离 个人眼粘液捐赠者。 隔离方法和 纯化将包括选择性溶剂萃取、亲和力和 离子交换色谱法、梯度超速离心法和 免疫学方法。 化学成分分析 分离物将包括 GLC 碳水化合物、氨基酸和 通过氨基酸分析确定氨基糖，通过薄层色谱分析确定脂质成分 和 GLC。 粘蛋白的结构分析将涉及氨基酸 肽核心测序以及 GLC、MS 和 NMR 光谱学与糖分析和免疫学相结合 寡糖链的测定。 （单克隆）抗体 粘蛋白、泪液前白蛋白和可能的去糖基化粘蛋白 将制备分离株。 个体物理特性研究 粘液成分，特别是粘蛋白，将包括 粘度、溶解度、分子的测量/测定 重量、表面活性和结构之间的结合方式 元素。 微量分析的最新进展和我们的经验 这些技术的应用将提供一个极好的 实现这些目标的机会。