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Published April 4, 2017 | Supplemental Material
Journal Article Open

Direct Intracellular Delivery of Cell Impermeable Probes of Protein Glycosylation Using Nanostraws

Abstract

Bioorthogonal chemistry is an effective tool for elucidating metabolic pathways and measuring cellular activity, yet its use is currently limited by the difficulty of getting probes past the cell membrane and into the cytoplasm, especially if more complex probes are desired. Here we present a simple and minimally perturbative technique to deliver functional probes of glycosylation into cells by using a nanostructured "nanostraw" delivery system. Nanostraws provide direct intracellular access to cells through fluid conduits that remain small enough to minimize cell perturbation. First, we demonstrate that our platform can deliver an unmodified azidosugar, N-azidoacetylmannosamine, into cells with similar effectiveness to a chemical modification strategy (peracetylation). We then show that the nanostraw platform enables direct delivery of an azidosugar modified with a charged uridine diphosphate group (UDP) that prevents intracellular penetration, thereby bypassing multiple enzymatic processing steps. By effectively removing the requirement for cell permeability from the probe, the nanostraws expand the toolbox of bioorthogonal probes that can be used to study biological processes on a single, easy-to-use platform.

Additional Information

© 2017 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim. Accepted manuscript online: 28 JAN 2017 12:50AM EST. Manuscript Accepted: 26 JAN 2017. Manuscript Revised: 26 JAN 2017. Manuscript Received: 21 DEC 2016. We acknowledge HFSP-RGP0048 and Bio-X Interdisciplinary Initiatives Program for support, and A.M.X. support through NSF and NDSEG graduate fellowships. We would like to acknowledge the Sarah Heilshorn Lab and the Stanford Neurofab facility for assisting with microscopy and cell culture access, the David Goldhaber-Gordon and Yi Cui Labs for use of equipment, and the Stanford Nanofabrication Facility and Stanford Nano Shared Facilities for fabrication and imaging.

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