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High-performance biosensors based on molecular orientation in enzyme thin films

基于酶薄膜分子取向的高性能生物传感器

基本信息

批准号：
11480252
负责人：
ANZAI Jun-ichi
金额：
$ 9.79万
依托单位：
Tohoku University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
1999
资助国家：
日本
起止时间：
1999 至 2002
项目状态：
已结题

项目摘要

Enzyme-containing thin films can be prepared by a layer-by-layer deposition of anionic or cationic polyelectrolyte and oppositely charged enzymes on a solid surface,through a electrostatic force of attraction. The thickness of the film depended significantly on the conditions of bathing solutions,such as polyelectrolyte and enzyme concentration,ionic strength,and pH. An alternate deposition of avidin and biotin-labeled enzymes also gave enzyme thin films through strong biological affinity between avidin and biotin. Avidin and enzyme formed a monomolecular layer in the multilayer film.The enzymes retained their catalytic activity in the film and thus it was possible to construct electrochemical enzyme biosensors by coating the film on the surface of electrodes.The biosensors thus prepared had an advantage that the size of output current can be regulated arbitrarily by changing the number of enzyme layers in the film. Glucose oxidase, lactate oxidase,uricase, and peroxidase were used as materials.Similarly, concanavalin A and glyco-enzymes can be used for constructing enzyme multilayer films,through affinity between concanavalin A and sugar. Non-glycosylated enzymes can also be used for this purpose after being modified with reactive sugar reagent. The concanavalin A-based enzyme thin films also exhibited catalytic activity.The layer-by-layer deposition technique developed in this study is applicable to the formation of thin film of antibody.The present technique would be useful for designing structure and function of biosensors.

含酶的薄膜可以通过阴离子或阳离子双酶和带相反电荷的酶通过静电吸引力在固体表面上逐层沉积来制备。膜的厚度取决于显着的条件下的浴溶液，如pH值和酶的浓度，离子强度，和pH值。交替沉积的亲和素和生物素标记的酶也通过亲和素和生物素之间的强大的生物亲和力酶薄膜。在多层膜中，亲和素和酶形成单分子层，酶在膜中保持其催化活性，因此，将膜涂敷在电极表面，可以构建电化学酶生物传感器，其优点是输出电流的大小可以通过改变膜中酶的层数来任意调节。以葡萄糖氧化酶、乳酸氧化酶、尿酸酶和过氧化物酶为材料，利用刀豆球蛋白A与糖的亲和性，以刀豆球蛋白A和糖酶为材料，构建酶多层膜。非糖基化的酶在用反应性糖试剂修饰后也可用于此目的。该方法可用于制备抗体薄膜，为生物传感器的结构和功能设计提供了依据。