Study of Membrane Domains with Coherent Raman Microscopy and Microspectroscopy

用相干拉曼显微镜和显微光谱学研究膜域

• 批准号: 0416785
• 负责人: Ji-Xin Cheng
• 金额: $ 47.9万
• 依托单位: Purdue University
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-08-01 至 2008-07-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=0416785&HistoricalAwards=false
• 关键词: Study; Membrane; Domains; Coherent; Raman

The objective of this project is to develop coherent anti-Stokes Raman scattering (CARS) microscopy to study membrane rafts. The PI will develop three-color CARS microscopy in which two images, one tuned to the peak and the other tuned to the dip of a CARS band, are acquired simultaneously. The difference of the two images will permit high-sensitivity vibration imaging of single membranes. The resonant CARS signal from the aliphatic C-D vibration will be used to determine the partition ratio of a specific lipid between the liquid ordered (rafts) and disordered domains. The PI will investigate whether rafts are formed by an interaction between cholesterol and lipid hydrocarbon chains or by hydrogen bonding between cholesterol and sphingomyelin. He will use the polarization sensitivity of CARS to determine the orientation of the CH2 groups of lipid hydrocarbon chains in a bilayer containing cholesterol. This method will be able to measure the ordering degree of lipids in rafts. The PI will also develop coherent Raman microspectroscopy for spectral characterization of lipid-lipid characterization in single membrane domains. The PI will directly test the hypothesis of lipid rafts in the plasma membrane of live cells. Aggregation of fluorophore labeled rafts has been observed in different cellular processes. He will examine if these macroscopic domains are rich in sphingomyelin and cholesterol using CARS microscopy. The non-resonant background that arises from water and other cellular organelles will be removed by using the three-color excitation scheme. This project provides an environment for interdisciplinary scientific training. Students involved in this research will need to combine the knowledge in chemistry, physics, and biology. The new imaging methods developed in this project will be incorporated in the PI's graduate level course of Biomedical Optics and undergraduate level course of Senior Engineering Design.

本计画的目的是发展相干反斯托克斯拉曼散射显微术来研究膜筏。PI将开发三色汽车显微镜，其中两个图像，一个调谐到峰值，另一个调谐到汽车波段的倾角，同时采集。两个图像的差异将允许单个膜的高灵敏度振动成像。来自脂肪族C-D振动的共振汽车信号将用于确定特定脂质在液体有序（筏）和无序结构域之间的分配比。 PI将研究筏是通过胆固醇和脂烃链之间的相互作用形成的，还是通过胆固醇和鞘磷脂之间的氢键形成的。他将使用汽车的偏振灵敏度来确定含有胆固醇的双层中类脂烃链的CH 2基团的方向。这种方法将能够测量筏中脂质的有序程度。PI还将开发相干拉曼显微光谱，用于单膜域中脂质-脂质表征的光谱表征。 PI将直接检验活细胞质膜中脂筏的假设。在不同的细胞过程中观察到荧光标记的筏的聚集。他将使用汽车显微镜检查这些宏观区域是否富含鞘磷脂和胆固醇。 由水和其他细胞器产生的非共振背景将通过使用三色激发方案被去除。该项目为跨学科科学培训提供了一个环境。参与这项研究的学生将需要联合收割机结合化学，物理和生物学的知识。 该项目中开发的新成像方法将纳入PI的生物医学光学研究生课程和高级工程设计本科课程中。