Nonlinear Imaging Methods for High-Throughput Protein and Drug Screening

项目摘要 越来越需要分析方法来确定和量化 低分子量药物化合物与生物膜的相互作用。的 脂质双层阵列的使用具有提供廉价和有效的高- 生产能力分析方法能够解决这些问题。有几个关键 如果脂质微阵列要达到成功的水平， 通过DNA微阵列分析获得。其中最主要的是需要一个 检测药物与脂质微阵列表面结合的非侵入性方法， 随后由于药物相互作用而引起的脂质基质的扰动。非线性 紫外-可见和频发生（UV-Vis SFG）和红外-可见和频发生（IR）技术， 可见光和频发生（IR-Vis SFG）成像可能对此有答案 问题.非线性成像的几个属性使其成为一种有前途的方法， 在微阵列上检测药物-膜相互作用，包括定量药物-膜相互作用的能力。 高光学分辨率和固有的表面特异性。的 关键的第一步，实施紫外-可见SFG和红外-可见SFG成像， 已经实现了脂质微阵列上药物缔合研究。 具体目标#1将侧重于开发紫外可见SFG，用于检测低浓度 分子量分子在平面支撑的脂质双层（PSLB）。使用UV- 维斯SFG用于测量膜的蛋白质吸附，其是 上一个赠款期的目标也将得到实现。非线性光谱 从几个模型蛋白质和药物靶点的反应将进行研究，试图 为了更充分地理解目标#1中控制其通过UV-Vis SFG检测的因素。 本文介绍了紫外-可见光谱SFG技术在离子通道抑制剂筛选中的应用。 探索目标#2药物对热致相的影响的检查 通过红外-可见SFG光谱和红外-可见光谱法研究了脂膜的相变和相行为。 目标#3中探索了维斯SFG高通量成像。这些研究旨在 展示了非线性成像方法与微图案化结合的实用性， 用于高通量药物筛选应用的流体脂质双层阵列。

Project Summary There is a growing need for analytical methods to identify and quantify the interaction of low molecular weight drug compounds with biological membranes. The use of lipid bilayer arrays has the potential to provide a cheap and efficient high- throughput analytical method capable of addressing these issues. There are several key issues which need to be resolved if lipid microarrays are to achieve the level of success obtained with DNA microarrays assays. Chief among these is the need for a noninvasive method to detect drug association to the lipid microarray surface and the subsequent perturbation of the lipid matrix due to drug interaction. The nonlinear techniques of ultraviolet-visible sum-frequency generation (UV-Vis SFG) and infrared- visible sum-frequency generation (IR-Vis SFG) imaging may hold the answer to this problem. Several attributes of nonlinear imaging make it a promising method for detecting drug-membrane interaction on microarrays, including the ability to quantify the detected signal, the high optical resolution and the inherent surface specificity. The crucial first steps to implementing UV-Vis SFG and IR-Vis SFG imaging for the investigation of drug association on lipid microarrays have already been achieved. Specific Aim #1 will focus on the development of UV-Vis SFG for the detection of low molecular weight molecules in planar supported lipid bilayers (PSLBs). The use of UV- Vis SFG for measuring protein adsorption to membranes, which is an extension of the goals of the previous grant period, will also be pursued. The nonlinear spectral response from several model protein and drug targets will be investigated in an attempt to more fully understand the factors controlling their detection by UV-Vis SFG in Aim #1. The application of UV-Vis SFG for the screening of potential ion channel inhibitors is explored in Aim #2. Examination of the influence of drugs on the thermotropic phase transition and phase behavior of lipid membranes by IR-Vis SFG spectroscopy and IR- Vis SFG high-throughput imaging is explored in Aim #3. These studies are designed to demonstrate the utility of nonlinear imaging methods in conjunction with micropatterned fluid lipid bilayer arrays for high-throughput drug screening applications.