XPS and SPR analysis of glycoarray surface density.

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.