Abstract
Polysaccharides have key biological functions and can be harnessed for therapeutic roles, such as the anticoagulant heparin. Their complexity—e.g., >100 monosaccharides with variety in linkage and branching structure—significantly complicates analysis compared to other biopolymers such as DNA and proteins. More, and improved, analysis tools have been called for, and here we demonstrate that solid-state silicon nitride nanopore sensors and tuned sensing conditions can be used to reliably detect native polysaccharides and enzymatic digestion products, differentiate between different polysaccharides in straightforward assays, provide new experimental insights into nanopore electrokinetics, and uncover polysaccharide properties. We show that nanopore sensing allows us to easily differentiate between a clinical heparin sample and one spiked with the contaminant that caused deaths in 2008 when its presence went undetected by conventional assays. The work reported here lays a foundation to further explore polysaccharide characterization and develop assays using thin-film solid-state nanopore sensors.
Highlights
Polysaccharides have key biological functions and can be harnessed for therapeutic roles, such as the anticoagulant heparin
Oligo- and polysaccharides are ubiquitous in nature, with a broad spectrum of roles that includes energy-storage and provision, structural building block, therapeutic function, and a vital part in biological recognition processes[1,2,3,4,5,6,7,8,9,10,11]
Conventional chemical analysis tools are frequently challenged by the daunting complexity of polysaccharide analysis:[12,13] identification of monomer composition (~120 naturally occurring monomers!) and sequence, monomer linkage types, stereochemistry, polymer length, and degree of polymer branching[13]
Summary
Polysaccharides have key biological functions and can be harnessed for therapeutic roles, such as the anticoagulant heparin. While solid-state nanopores in thin (~10 nm) membranes have been often portrayed as the preeminent nanopore platform, their use to profile classes of molecules beyond DNA and proteins is in its infancy These nanopores can be size-tuned[44] to match analyte dimensions (especially relevant for branched polysaccharides), and when fabricated from conventional nanofabrication materials such as silicon nitride (SiNx)[45,46], offer resistance to chemical and mechanical insult alongside low barriers to large-scale manufacturing and device integration. This exceptional charge density couples with the demonstrated difficulty, by other methods, of detecting the negatively charged OSCS (molecular weight ~17 kDa51) contaminant in a heparin sample[14,15,16,17] to make the analysis of heparin (~16 kDa) and OSCS by nanopore a compelling experimental test with clinical relevance
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