Printed protein microarrays on unmodified plastic substrates.
Printed protein microarrays on unmodified plastic substrates.
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DOI：
10.1016/j.aca.2010.05.008
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发表时间：
2010-06
影响因子：
6.2
通讯作者：
M. Moschallski;J. Baader;O. Prucker;J. Rühe
中科院分区：
文献类型：
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作者：
M. Moschallski;J. Baader;O. Prucker;J. Rühe
蛋白质微阵列产生的一个关键挑战是将功能捕获探针蛋白质固定在芯片表面。在这里，提出了一种新的概念，即在未经修饰的塑料基质上一步生产蛋白质微阵列。它的基础是聚合物/蛋白质混合物的印刷和所获得的微结构的光化学附着到塑料芯片表面。在光化学过程中，三个反应同时发生：聚合物转变为水凝胶点，形成的凝胶与底物共价结合，蛋白质共价固定到三维水凝胶支架上。作为例子，我们使用抗牛血清白蛋白作为蛋白质(anti-BSA)和以聚二甲基丙烯酰胺为支架的水膨胀聚合物网络，并以二苯甲酮为交联剂进行光化学交联。在一系列微阵列实验中，通过在水凝胶中掺入荧光标记的抗BSA来确定固定化蛋白质的探针密度。在一个典型的实验中，固定化探针的数量被确定为每个点4×109个蛋白质分子。在其他实验中，将微阵列与荧光标记的BSA接触。在这种分析中，获得了超过200的信噪比，每个斑点大约结合了9×107个抗原分子。这表明，以一种非常简单的方式可以实现每个点具有大量探针的微阵列，并且固定在印刷水凝胶中的抗体仍然可访问并保持其功能。
A key challenge for the generation of protein microarrays is the immobilization of functional capture probe proteins at the chip surfaces. Here, a new concept for a single step production of protein microarrays to unmodified plastic substrates is presented. It is based on the printing of polymer/protein mixtures and the photochemical attachment of the obtained microstructures to the plastic chip surfaces. In the photochemical process three reactions occur simultaneously: transformation of the polymer into hydrogel dots, covalent binding of the forming gel to the substrate, and covalent immobilization of the proteins to the three-dimensional hydrogel scaffold. As an example we use anti-bovine serum albumin as a protein (anti-BSA) and a water swellable polymer network based on polydimethylacrylamide as a scaffold, which is photochemically crosslinked using benzophenone as a crosslinking agent. In one series of microarray experiments the probe density of the immobilized proteins was determined by incorporating fluorescence-labeled anti-BSA in the hydrogels. In a typical experiment, the number of immobilized probes was determined to 4×109protein molecules per spot. In other experiments, the microarrays were brought into contact with fluorescently labeled BSA. In such analyses signal-to-noise values of more than 200 were obtained and about 9×107antigen molecules were bound per spot. This demonstrates that in a very simple way microarrays with large amount of probes per spot can be realized and that antibodies immobilized in the printed hydrogels remain accessible and retain their functionality.