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DOE OSTI · 2565502

DNA-PAINT Imaging with Hydrogel Imprinting and Clearing

Stein, Johannes [Harvard Medical School, Boston, MA (United States)] (ORCID:0000000213351120)·Magni, Lorenzo [Wyss Institute of Biologically Inspired Engineering, Boston, MA (United States); Harvard Medical School, Boston, MA (United States)]·Church, George M. [Wyss Institute of Biologically Inspired Engineering, Boston, MA (United States); Harvard Medical School, Boston, MA (United States)] (ORCID:0000000335352076)
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Abstract

Hydrogel-embedding is a versatile technique in fluorescence microscopy, offering stabilization, optical clearing, and the physical expansion of biological specimens. DNA-PAINT is a super-resolution microscopy approach based on the diffusion and transient binding of fluorescently labeled oligos, but its feasibility in hydrogels has not yet been explored. In this study, we demonstrate that polyacrylamide hydrogels support sufficient diffusion for effective DNA-PAINT imaging. Using acrydite-anchored oligonucleotides imprinted from patterned DNA origami nanostructures and microtubule filaments in fixed cells, we find that hydrogel embedding preserves docking strand positioning at the nanoscale. Sample clearing via protease treatment had minor structural effects on the microtubule structure and enhanced diffusion and accessibility to hydrogel-imprinted docking strands. Our work demonstrates promising potential for diffusion and binding-based fluorescence imaging applications in hydrogel-embedded samples.

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DNA origamiDNA-PAINTDiffusionEmbeddingHydrogel embeddingHydrogelsMatrix imprinting and clearingPeptides and proteinsPolyacrylamide gelsSingle-molecule localization microscopySuper-resolution microscopy

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Stein, Johannes [Harvard Medical School, Boston, MA (United States)] (ORCID:0000000213351120), Magni, Lorenzo [Wyss Institute of Biologically Inspired Engineering, Boston, MA (United States); Harvard Medical School, Boston, MA (United States)], Church, George M. [Wyss Institute of Biologically Inspired Engineering, Boston, MA (United States); Harvard Medical School, Boston, MA (United States)] (ORCID:0000000335352076). 2025-05-09. DNA-PAINT Imaging with Hydrogel Imprinting and Clearing. https://doi.org/10.1021/acssensors.5c00616

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Transport of Delocalized Excitons through DNA-Based Molecular Photonic Wires

Molecular photonic wires conduct electronic energy via their rapid transport properties. In photosynthesis, nature achieves efficient transport across large distances using delocalized excitons, generated by strong excitonic coupling between chromophores. How, or even whether, delocalization facilitates long-distance energy transport in synthetic systems has been challenging to experimentally test and optimize. Thus, far, studies have been limited to strongly coupled, heterogeneous chromophore aggregates or weakly coupled chromophore monomers. Here, in this work, we employed DNA nanostructures to engineer molecular photonic wires constructed from a series of excitonically coupled indocarbocyanine chromophores─achieving the intermediate and strong coupling regimes. Using time-resolved fluorescence spectroscopy and complementary simulations, we demonstrated that an intermediate intermolecular electronic coupling (∼ k B T ) enables up to 40% faster exciton transport as compared to strongly coupled chromophores. The delocalized excitons generated in the intermediate coupling regime exhibited properties conducive to rapid diffusivity, similar to their monomeric counterparts. Thus, intermediate excitonic coupling, analogous to natural systems, achieves long-distance exciton transport with the high chromophore density required for energy capture.

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