Analysis of protein adsorption and binding at biosensor polymer interfaces using X-ray photon spectroscopy and scanning electrochemical microscopy
Analysis of protein adsorption and binding at biosensor polymer interfaces using X-ray photon spectroscopy and scanning electrochemical microscopy
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DOI：
10.1021/ac0261653
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发表时间：
2003-06-01
影响因子：
7.4
通讯作者：
Cooper, JM
中科院分区：
文献类型：
--
作者：
Glidle, A;Yasukawa, T;Cooper, JM
我们描述了一种方法，基于X射线光电子能谱（XPS）测量，以评估蛋白质吸附或结合在各种不同的yTAS和生物传感器接口的程度。支持这种方法的是通过使用先前为放射性示踪剂研究开发的协议用含碘或含溴基序标记蛋白质分子。使用这种方法，我们已经研究了各种改性的电沉积聚合物界面以及其他材料中使用的pTAS器件制造的吸附和结合性能。使用聚丙二醇（PPG）链修饰的聚合物界面，我们的研究结果表明，至少需要一个类似于30个单体单元的链才能抑制浓缩蛋白质溶液的非特异性吸附。XPS方法还用于探测抗生物素蛋白及其酶缀合物与生物素化的和混合的生物素/PPG修饰的聚合物界面的特异性结合。在一个实施例中，使用竞争性结合，确定过氧化物酶-链霉亲和素缀合物与生物素化的改性聚合物界面的结合模式主要是经由链霉亲和素部分（与经由酶缀合物的非特异性结合相反）。XPS评估非特异性和特异性过氧化物酶-链霉亲和素固定在各种功能化聚合物上指导了生物传感muTAS装置中的功能化叉指电极的设计和制造。随后使用扫描电化学显微镜（SECM）对该装置进行表征，证实了先前从宏观电极上的XPS测量推断的吸附和结合。
We describe a method, based on X-ray photoelectron spectroscopy (XPS) measurements, to assess the extent of protein adsorption or binding on a variety of different yTAS and biosensor interfaces. Underpinning this method is the labeling of protein molecules with either iodine- or bromine-containing motifs by using protocols previously developed for radiotracer studies. Using this method, we have examined the adsorption and binding properties of a variety of modified electrodeposited polymer interfaces as well as other materials used in pTAS device fabrication. Using polymer interfaces modified with poly(propylene glycol) (PPG) chains, our results indicate that a chain of at least similar to30 monomer units is required to inhibit nonspecific adsorption from concentrated protein solutions. The XPS methodology was also used to probe specific binding of avidins and enzyme conjugates thereof to biotinylated and mixed biotin/PPG-modified polymer interfaces. In one example, using competitive binding, it was established that the mode of binding of a peroxidase-streptavidin conjugate to a biotinylated modified polymer interface was primarily via the streptavidin moiety (as opposed to nonspecific binding via the enzyme conjugate). XPS evaluation of nonspecific and specific peroxidase-streptavidin immobilization on various functionalized polymers has guided the design and fabrication of functionalized interdigitated electrodes in a biosensing muTAS device. Subsequent characterization of this device using scanning electrochemical microscopy (SECM) corroborated the adsorption and binding previously inferred from XPS measurements on macroscale electrodes.