Creating spatially addressed arrays of planar supported fluid phospholipid membranes
Creating spatially addressed arrays of planar supported fluid phospholipid membranes
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DOI：
10.1021/ja991453a
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发表时间：
1999-09-08
影响因子：
15
通讯作者：
Yang, TL
中科院分区：
文献类型：
--
作者：
Cremer, PS;Yang, TL
使用平面载体来呈现大量的空间寻址分子阵列是创建组合文库的最强大和最通用的方法之一。1-3这些系统开始为生物和化学科学中的新一代快速筛查分析和传感器设备奠定基础。将这种方法扩展到含有多肽、受体和完整膜蛋白的支撑型磷脂双层膜是一个特别有价值的目标，因为这些系统能够模拟天然细胞表面的许多特性。4然而，在平面支撑物上处理生物膜模拟存在独特的挑战，因为在许多情况下必须保持生物膜的二维流动性，才能使其正常发挥作用。5-7双层沉积过程必须在水环境中进行，整个系统必须继续淹没在水下，以保存平面支撑结构。由于这种物理限制以及生物膜材料固有的复杂性，传统的技术，如将多肽或DNA序列寻址到固体载体上的光引导合成，本身就很难应用。1因此，我们采用了另一种方法，基于将介观数量的水溶液沉积到光刻图案的亲水表面孔板上，8然后将整个衬底浸入缓冲液中。这是一种通用和灵活的方法，用于将化学上不同的磷脂膜引导到单独可寻址的表面扇区。以前的研究表明，图案化的表面允许将一个液体脂双层与另一个液体脂双层隔开。单个膜内的10个分子可以在单个隔板的范围内自由移动，但不能跨越到邻近区域。在这里介绍的实验中，使用标准光刻技术将平面硼硅酸盐衬底分割成微米大小的亲水性盒子阵列。图案化是通过由方框阵列组成的光刻掩模将表面暴露在紫外光下实现的。显影图案和清洁衬底形成了良好的亲水性玻璃板，在其上放置了皮升大小的脂质体溶液液滴(图1)。脂质体为磷脂的小单层囊泡，在pH 7.0,100 mM的磷酸二氢钠缓冲溶液中，浓度为1 mg/mL。
The use of planar supports for presenting large arrays of spatially addressed molecules is one of the most powerful and versatile methods for creating combinatorial libraries. 1-3 These systems are starting to form the basis for a new generation of rapid screening assays and sensor devices in the biological and chemical sciences. Extending this approach to supported phospholipid bilayer membranes containing peptides, receptors, and integral membrane proteins is an especially valuable goal because of the ability of these systems to mimic many of the properties of native cell surfaces. 4 Addressing biomembrane mimics on planar supports, however, presents unique challenges, as the twodimensional fluidity of the biomembrane must be preserved in many cases for it to function properly. 5-7 The bilayer deposition process must take place in an aqueous environment, and the entire system must continue to remain submerged under water to preserve the planar supported structure. Because of this physical constraint as well as the inherent complexities of biomembrane materials, traditional technologies such as light-directed synthesis for addressing peptide or DNA sequences onto solid supports are inherently difficult to apply. 1 We have, therefore, employed an alternate approach based upon depositing mesoscopic quantities of aqueous solution onto lithographically patterned hydrophilic surface well plates, 8 followed by the immersion of the entire substrate into buffer. This is a general and flexible method for directing chemically distinct phospholipid membranes into individually addressable surface sectors. Previous studies have shown that patterned surfaces allow partitioning of one fluid lipid bilayer from the next. 9, 10 Molecules within an individual membrane are free to move within the confines of a single partition but do not cross over to a neighboring region. In the experiments presented here, planar borosilicate substrates were partitioned into arrays of micrometer-sized hydrophilic boxes using standard photolithography. Patterning was achieved by exposing the surface to ultraviolet light through a lithographic mask consisting of an array of square boxes. Developing the pattern and cleaning the substrate formed well plates of hydrophilic glass onto which picoliter-sized droplets of liposome solution were placed (Figure 1). The liposomes, which were small unilamellar vesicles (SUVs) of phospholipids, were present at 1 mg/mL concentration in a pH 7.0, 100 mM sodium phosphate buffer solution.