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Reversible Functionalization of Clickable Polyacrylamide Gels with Protein and Graft Copolymers.

ABSTRACT: Modular strategies to fabricate gels with tailorable chemical functionalities are relevant to applications spanning from biomedicine to analytical chemistry. Here, the properties of clickable poly(acrylamide-co-propargyl acrylate) (pAPA) hydrogels are modified via sequential in-gel copper-catalyzed azide-alkyne cycloaddition (CuAAC) reactions. Under optimized conditions, each in-gel CuAAC reaction proceeds with rate constants of ~0.003 s^-1, ensuring uniform modifications for gels < 200 μm thick. Using the modular functionalization approach and a cleavable disulfide linker, pAPA gels were modified with benzophenone and acrylate groups. Benzophenone groups allow gel functionalization with unmodified proteins using photoactivation. Acrylate groups enabled copolymer grafting onto the gels. To release the functionalized unit, pAPA gels were treated with disulfide reducing agents, which triggered ~50 % release of immobilized protein and grafted copolymers. The molecular mass of grafted copolymers (~6.2 kDa) was estimated by monitoring the release process, expanding the tools available to characterize copolymers grafted onto hydrogels. Investigation of the efficiency of in-gel CuAAC reactions revealed limitations of the sequential modification approach, as well as guidelines to convert a pAPA gel with a single functional group into a gel with three distinct functionalities. Taken together, we see this modular framework to engineer multifunctional hydrogels as benefiting applications of hydrogels in drug delivery, tissue engineering, and separation science.

SUBMITTER: Neira HD

PROVIDER: S-EPMC7942169 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Hydrogels are important structural and operative components of microfluidic systems, finding diverse utility in biological sample preparation and interrogation. One inherent challenge for integrating hydrogels into microfluidic tools is thermodynamic molecular partitioning, which reduces the in-gel concentration of molecular solutes (e.g., biomolecular regents), as compared to the solute concentration in an applied solution. Consequently, biomolecular reagent access to in-gel scaffolded biological samples (e.g., encapsulated cells, microbial cultures, target analytes) is adversely impacted in hydrogels. Further, biomolecular reagents are typically introduced to the hydrogel via diffusion. This passive process requires long incubation periods compared to active biomolecular delivery techniques. Electrotransfer is an active technique used in Western blots and other gel-based immunoassays that overcomes limitations of size exclusion (increasing the total probe mass delivered into gel) and expedites probe delivery, even in millimeter-thick slab gels. While compatible with conventional slab gels, electrotransfer has not been adapted to thin gels (50-250 μm thick), which are of great interest as components of open microfluidic devices (vs enclosed microchannel-based devices). Mechanically delicate, thin gels are often mounted on rigid support substrates (glass, plastic) that are electrically insulating. Consequently, to adapt electrotransfer to thin-gel devices, we replace rigid insulating support substrates with novel, mechanically robust, yet electrically conductive nanoporous membranes. We describe grafting nanoporous membranes to thin-polyacrylamide-gel layers via silanization, characterize the electrical conductivity of silane-treated nanoporous membranes, and report the dependence of in-gel immunoprobe concentration on transfer duration for passive diffusion and active electrotransfer. Alternative microdevice component layers─including the mechanically robust, electrically conductive nanoporous membranes reported here─provide new functionality for integration into an increasing array of open microfluidic systems.

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Reversible Functionalization of Clickable Polyacrylamide Gels with Protein and Graft Copolymers.

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