Department of Chemistry and the Carolina Center for Genome Science, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina 27599-3290, USA.
Langmuir. 2010 Jun 15;26(12):9835-41. doi: 10.1021/la1001545.
We report a new, quantitative methodology to pattern and present ligands from planar, supported, fluid lipid bilayers. By combining microfluidic lithography (microFL) with an electroactive, chemoselective interfacial reaction strategy, a number of ligands as well as protein concanavalin A were immobilized in lipid microarrays. Electroactive vesicles were generated after the spontaneous insertion of hydroquinone-tethered alkane (H(2)Q) into egg palmitoyl-oleoyl phosphatidylcholine (egg-POPC), followed by subsequent fusion to a siloxane-terminated self-assembled monolayer (SAM) on gold. An advantage of the H(2)Q system is that it can be electrochemically oxidized to the corresponding quinone (Q), followed by rapid chemoselective conjugation with oxyamine-functionalized (RONH(2)) ligands. The oxime product is also electroactive, and the reaction can be monitored and the amount of ligand bound can be quantified by electrochemistry. The bilayers were characterized by electrochemistry, fluorescence microscopy, and ellipsometry and were determined to be fluid by fluorescence recovery after photobleaching (FRAP). This strategy provides a synergistic method to pattern and present a number of ligands or biomolecules from the bilayer surface for the evaluation of enzyme or protein binding to biomembranes.
我们报告了一种新的、定量的方法，可以对平面支撑的流体脂质双层中的配体进行图案化和呈现。通过将微流控光刻（microFL）与电化学生物选择性界面反应策略相结合，可以将多种配体以及蛋白伴刀豆球蛋白 A 固定在脂质微阵列中。在将氢醌键合烷烃（H（2）Q）自发插入蛋黄棕榈酰-油酰磷脂酰胆碱（egg-POPC）中之后，生成电活性囊泡，随后融合到金上的硅氧烷封端自组装单层（SAM）上。H（2）Q 体系的一个优点是它可以电化学氧化为相应的醌（Q），然后与氧基胺功能化（RONH（2））配体快速进行化学选择性缀合。肟产物也是电活性的，并且可以通过电化学监测和定量结合到配体的量来定量。通过电化学、荧光显微镜和椭圆光度法对双层进行了表征，并且通过光漂白后荧光恢复（FRAP）确定其为流体。该策略提供了一种协同方法，可以从双层表面对多种配体或生物分子进行图案化和呈现，以评估酶或蛋白质与生物膜的结合。
