Department of Chemistry and Biochemistry and California NanoSystems Institute, University of California, Los Angeles , 607 Charles E. Young Drive East, Los Angeles, California 90095-1569, United States.
Langmuir. 2016 Apr 26;32(16):4043-51. doi: 10.1021/acs.langmuir.6b00560. Epub 2016 Apr 14.
Electron beam (e-beam) lithography was employed to prepare one protein immobilized hydrogel encapsulated inside another by first fabricating protein-reactive hydrogels of orthogonal reactivity and subsequently conjugating the biomolecules. Exposure of thin films of eight arm star poly(ethylene glycol) (PEG) functionalized with biotin (Biotin-PEG), alkyne (Alkyne-PEG) or aminooxy (AO-PEG) end-groups to e-beam radiation resulted in cross-linked hydrogels with the respective functionality. It was determined via confocal microscopy that a nominal size exclusion effect exists for streptavidin immobilized on Biotin-PEG hydrogels of feature sizes ranging from 5 to 40 μm. AO-PEG was subsequently patterned as an encapsulated core inside a contiguous outer shell of Biotin-PEG. Similarly, Alkyne-PEG was patterned as a core inside an AO-PEG shell. The hydrogel reactive end-groups were conjugated to dyes or proteins of complementary reactivity, and the three-dimensional (3-D) spatial orientation was determined for both configurations using confocal microscopy. The enzyme glucose oxidase (GOX) was immobilized in the core of the encapsulated Alkyne-PEG core/ AO-PEG shell architecture, and horseradish peroxidase (HRP) was conjugated to the shell periphery. Bioactivity for the HRP-GOX enzyme pair was observed in this encapsulated configuration by demonstrating that the enzyme pair was capable of enzyme cascade reactions.
电子束（e-beam）光刻被用于通过首先制备具有正交反应性的蛋白质反应性水凝胶，然后缀合生物分子，来制备一种蛋白质固定的水凝胶包裹在另一种水凝胶内部。将带有生物素（Biotin-PEG）、炔基（Alkyne-PEG）或氨氧基（AO-PEG）末端基团的八臂星形聚（乙二醇）（PEG）的薄膜暴露于电子束辐射下，导致具有相应功能的交联水凝胶。通过共聚焦显微镜确定，对于固定在特征尺寸为 5 至 40μm 的 Biotin-PEG 水凝胶上的链霉亲和素，存在标称的尺寸排除效应。AO-PEG 随后被图案化为 Biotin-PEG 连续外壳内的封装核。同样，Alkyne-PEG 被图案化为 AO-PEG 壳内的核。水凝胶反应性末端基团与具有互补反应性的染料或蛋白质缀合，并且使用共聚焦显微镜确定了这两种构型的三维（3-D）空间取向。将葡萄糖氧化酶（GOX）固定在封装的 Alkyne-PEG 核/ AO-PEG 壳结构的核中，并将辣根过氧化物酶（HRP）缀合到壳的外围。通过证明该酶对可以进行酶级联反应，观察到这种封装构型中的 HRP-GOX 酶对的生物活性。
