Abstract
Covalent modification of surfaces with carbohydrates (glycans) is a prerequisite for a variety of glycomics-based biomedical applications, including functional biomaterials, glycoarrays, and glycan-based biosensors. The chemistry of glycan immobilization plays an essential role in the bioavailability and function of the surface bound carbohydrate moiety. However, the scarcity of analytical methods to characterize carbohydrate-modified surfaces complicates efforts to optimize glycan surface chemistries for specific applications. Time-of-Flight Secondary Ion Mass Spectrometry (ToF-SIMS) is a surface sensitive technique suited for probing molecular composition at the biomaterial interface. Expanding ToF-SIMS analysis to interrogate carbohydrate-modified materials would increase our understanding of glycan surface chemistries and advance novel tools in the nascent field of glycomics. In this study, a printed glycan microarray surface was fabricated and subsequently characterized by ToF-SIMS imaging analysis. A multivariate technique based on principal component analysis (PCA) was used to analyze the ToF-SIMS dataset and reconstruct ToF-SIMS images of functionalized surfaces. These images reveal chemical species related to the immobilized glycan, underlying glycan-reactive chemistries, gold substrates, and outside contaminants. Printed glycoarray elements (spots) were also interrogated to resolve the spatial distribution and spot homogeneity of immobilized glycan. The bioavailability of the surface-bound glycan was validated using a specific carbohydrate-binding protein (lectin) as characterized by Surface Plasmon Resonance Imaging (SPRi). Our results demonstrate that ToF-SIMS is capable of characterizing chemical features of carbohydrate-modified surfaces and, when complemented with SPRi, can play an enabling role in optimizing glycan microarray fabrication and performance.
Highlights
Advances in synthetic carbohydrate chemistry have enabled the creation of complex carbohydratemodified surfaces for biomedical and glycobiology research [1,2,3,4]
Carbohydrate microarrays were fabricated by printing a diaminopyridine-modified glycan (GDAP, see Scheme 1 in the Experimental Section) onto a model surface consisting of an NHS-activated oligoethylene glycol self-assembled monolayer (OEG/self-assembled monolayers (SAMs))
In order for the carbohydrate microarray to realize its full potential as a tool for biomedical research and drug discovery, the microarray’s biointerface must be rigorously interrogated and optimized to account for the effects of surface inhomogeneity, printing artifacts, and adventitious contaminants
Summary
Advances in synthetic carbohydrate chemistry have enabled the creation of complex carbohydratemodified surfaces for biomedical and glycobiology research [1,2,3,4]. Arrays were evaluated against a simple panel of fluorescence-labeled carbohydrate-binding proteins with known binding specificities, but this method did not have well-established protocols for translating these results into quantifiable information on array surface chemistry and performance This meant that until recently the quality of the printed glycan array was largely an unknown entity. Carbohydrate microarrays were fabricated by printing a diaminopyridine-modified glycan (GDAP, see Scheme 1 in the Experimental Section) onto a model surface consisting of an NHS-activated oligoethylene glycol self-assembled monolayer (OEG/SAM) For this preliminary study, self-assembled monolayers (SAMs) were selected over commercial array substrates for their reliability, simple chemistries and extensive surface characterization by XPS, ToF-SIMs and SPR. Peaks of interest were determined using PCA and the distribution of select molecular species was visualized and compared to SPRi bioactivity This combined chemical- and biological-imaging approach provides insight into the factors that affect carbohydrate microarray performance
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