Kalyankar Nikhil D, Sharma Manoj K, Vaidya Shyam V, Calhoun David, Maldarelli Charles, Couzis Alexander, Gilchrist Lane
Department of Chemical Engineering, City University of New York, New York, USA.
Langmuir. 2006 Jun 6;22(12):5403-11. doi: 10.1021/la0602719.
Here, we describe a protocol to bind individual, intact phospholipid bilayer liposomes, which are on the order of 1 microm in diameter, in microwells etched in a regular array on a silicon oxide substrate. The diameter of the wells is on the order of the liposome diameter, so only one liposome is located in each well. The background of the silicon oxide surface is functionalized with a PEG oligomer using the contact printing of a PEG silane to present a surface that resists the adsorption of proteins, lipid material, and liposomes. The interiors of the wells are functionalized with an aminosilane to facilitate the conjugation of biotin, which is then bound to Neutravidin. The avidin-coated well interiors bind the liposomes whose surfaces contain biotinylated lipids. The specific binding of the liposomes to the surface using the biotin-avidin linkage, together with the resistant nature of the background and the physical confinement of the wells, allows the liposomes to remain intact and to not unravel, rupture, and fuse onto the surface. We demonstrate this intact arraying using confocal laser scanning microscopy of fluorophores specifically tagging the microwells, the lipid bilayer, and the aqueous interior of the liposome.
在此，我们描述了一种将直径约为1微米的单个完整磷脂双层脂质体结合到在氧化硅基板上以规则阵列蚀刻的微孔中的方法。孔的直径与脂质体直径相当，因此每个孔中仅定位一个脂质体。使用聚乙二醇硅烷的接触印刷法对氧化硅表面进行功能化处理，以呈现出一种能抵抗蛋白质、脂质材料和脂质体吸附的表面。孔的内部用氨基硅烷进行功能化处理，以促进生物素的缀合，然后生物素与中性抗生物素蛋白结合。涂有抗生物素蛋白的孔内部结合了表面含有生物素化脂质的脂质体。利用生物素 - 抗生物素蛋白连接将脂质体特异性结合到表面，再加上背景的抗性和孔的物理限制，使得脂质体能够保持完整，不会在表面展开、破裂或融合。我们使用共聚焦激光扫描显微镜对特异性标记微孔、脂质双层和脂质体水相内部的荧光团进行观察，展示了这种完整的阵列排列。
