Dhayal Marshal, Ratner Daniel M
Department of Bioengineering, University of Washington, Seattle, Washington 98195, USA.
Langmuir. 2009 Feb 17;25(4):2181-7. doi: 10.1021/la8031122.
Despite the fact that the carbohydrate microarray has seen increasing use within the field of glycobiology, the surface chemistry of the glycoarray remains largely unexplored. Motivated by the need to develop surface analytical techniques to characterize carbohydrate-modified surfaces, we developed a quantitative X-ray photoelectron spectroscopy (XPS) and surface plasmon resonance imaging (SPR imaging) method to study glycan biosensors. We performed a comparative analysis on the relative coverage of mixed self-assembled monolayers (SAMs) on gold, consisting of a thiol-functionalized trimannoside (Manalpha1,2Manalpha1,2Manalpha-OEG-SH) at varying concentrations (0-100%) mixed separately with two thiol-containing polyethylene glycol oligomers. XPS C1s core level analysis was used to identify the O-C-O functionality unique to the carbohydrate acetal moiety and to separate and quantify the relative coverage of sugar in carbohydrate/OEG mixed SAMs. XPS spectra of the mixed SAMs demonstrated a proportional increase in the acetal signature of the glycan with increasing sugar concentration. To relate surface glycan density with biological function, we carried out a kinetic analysis of Concanavalin A (ConA) binding to SAMs of varying densities of carbohydrate using SPR imaging. We observed protein binding that was highly dependent on both glycan density and the nature of the OEG-thiol used in the mixed self-assembly. These results illustrate the utility of surface analytical techniques such as XPS and SPR in carbohydrate biosensor characterization and optimization.
尽管碳水化合物微阵列在糖生物学领域的应用日益广泛，但糖芯片的表面化学仍 largely未被探索。出于开发用于表征碳水化合物修饰表面的表面分析技术的需要，我们开发了一种定量X射线光电子能谱（XPS）和表面等离子体共振成像（SPR成像）方法来研究聚糖生物传感器。我们对金表面混合自组装单分子层（SAMs）的相对覆盖率进行了比较分析，该混合自组装单分子层由不同浓度（0 - 100%）的硫醇官能化三甘露糖苷（Manα1,2Manα1,2Manα - OEG - SH）分别与两种含硫醇的聚乙二醇低聚物混合组成。XPS C1s核心能级分析用于识别碳水化合物缩醛部分特有的O - C - O官能团，并分离和量化碳水化合物/OEG混合SAMs中糖的相对覆盖率。混合SAMs的XPS光谱表明，随着糖浓度的增加，聚糖的缩醛特征呈比例增加。为了将表面聚糖密度与生物学功能联系起来，我们使用SPR成像对伴刀豆球蛋白A（ConA）与不同碳水化合物密度的SAMs的结合进行了动力学分析。我们观察到蛋白质结合高度依赖于聚糖密度和混合自组装中使用的OEG - 硫醇的性质。这些结果说明了XPS和SPR等表面分析技术在碳水化合物生物传感器表征和优化中的实用性。
