IN SITU MASS SPECTRAL ANALYSIS OF ELECTROPHORETOGRAMS

电泳图的原位质谱分析

• 批准号: 3290461
• 负责人: KENNETH L BUSCH
• 金额: $ 5.9万
• 依托单位: TRUSTEES OF INDIANA UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1986
• 资助国家: 美国
• 起止时间: 1986-07-01 至 1989-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3290461
• 关键词: analytical method; automated data processing; biochemistry; carnitine; electrophoresis; hemoglobin; mass spectrometry; paper chromatography; peptides; thin layer chromatography

Biochemical studies make extensive use of static chromatographies such as high resolution two dimensional paper and thin layer chromatography and electrophoresis. The information available from such techniques is limited by the relatively poor performance of the detection systems traditionally used. This proposal supports the development of a mass spectrometric system which can analyze separated biomolecules in situ, with a particular focus on the analysis of peptide mixtures separated by high performance two dimensional electrophoresis. With information available from the mass spectrum, the identity and homogeneity of the separated components can be established without loss of the integrity of the chromatogram. In favorable instances, the compounds can be recovered after analysis for further characterization or processing. In this new instrument, intact chromatograms are placed under vacuum, and components encapsulated in the matrix are brought into focus by movement of the chromatogram in the xy development plane. A secondary ion mass spectrometer with a highly focused primary ion beam sputters material directly from the chromatogram into a quadrupole mass analyzer. A spatial resolution of 10 microns in both x and y can increase the effective resolution of poorly separated compounds by processing of the mass spectral data. A feedback system automatically maximizes the efficiency of data collection by increasing spatial resolution as compounds are brought into the instrument focus. Initial applications will be to the analysis of carnitine-based samples separated by thin layer chromatography, to sickle cell hemoglobin peptides separated by electrophoresis, and to a new experiment, selected sequence monitoring, which pinpoints peptides containing a specific sequence of amino acids from within a mixture separated by two dimensional chromatography. These initial experiments document the performance of the new system in problems which have significant biochemical interest. By the end of the proposed period, an automated analysis system will have been developed which can be used for general analysis of complex mixtures separated by two dimensional electrophoresis.

生物化学研究广泛使用静态色谱法，如 高分辨率二维纸和薄层色谱， 电泳 从这些技术中获得的信息是有限的 由于传统上检测系统的相对较差的性能 采用 这一建议支持质谱的发展， 一种可以原位分析分离的生物分子的系统， 专注于分析肽混合物分离的高性能二 双向电泳 从大众那里得到的信息 通过光谱，可以确定分离组分的同一性和均匀性。 在不损失色谱图完整性的情况下建立。 在 在有利的情况下，化合物可以在分析后回收， 进一步表征或加工。 在这个新仪器中， 将色谱图置于真空下，并将包封在 通过色谱图在xy方向上的移动， 发展平面 一种高聚焦二次离子质谱仪， 一次离子束直接从色谱图中分离出物质， 四极质量分析器 在x和x方向上的空间分辨率均为10微米， y可以通过以下方式提高分离效果差的化合物的有效分离度： 质谱数据的处理。 反馈系统自动 通过增加空间分辨率，最大限度地提高数据收集效率 当化合物被带入仪器焦点时， 初始 应用将是分析肉毒碱为基础的样品分离 通过薄层色谱法，将镰状细胞血红蛋白肽分离出来 通过电泳，和一个新的实验，选择序列监测， 它可以精确定位含有特定氨基酸序列的肽， 在通过二维色谱分离的混合物中。 这些 初步的实验证明了新系统在问题中的性能 其具有重要的生物化学意义。 截至2010年12月31日， 在此期间，将开发一个自动分析系统， 用于二维分离的复杂混合物的一般分析 电泳