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bioRxiv · 10.1101/2023.10.31.564845

High-throughput, label-free detection of DNA origami in single-cell suspensions using origamiFISH-Flow

Alexander, S.·Wang, W. X.·Tseng, C.-Y.·Douglas, T. R.·Chou, L. Y. T.
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Abstract

Structural DNA nanotechnology enables custom fabrication of nanoscale devices and promises diverse biological applications. However, the effects of design on DNA nanostructure (DN)-cell interactions in vitro and in vivo are not yet well-characterized. origamiFISH is a recently developed technique for imaging DNs in cells and tissues. Compared to the use of fluorescent tags, origamiFISH offers label-free and structure-agnostic detection of DNs with significantly improved sensitivity. Here, we extend the origamiFISH technique to quantifying DNs in single-cell suspensions, including nonadherent cells such as subsets of immune cells, via readout by flow cytometry. This method, referred to as origamiFISH-Flow, is high-throughput (e.g., 10,000 cells per second) and compatible with immunostaining for concurrent cell-type and -state characterization. We demonstrate that origamiFISH-Flow enhances signal-to-noise ratio by up to 20-fold compared to dye labeling approaches, leading to the capture of >25-fold more DN+ cells at low, single-picomolar DN uptake concentrations. We additionally show the use of origamiFISH-Flow to profile cell-type and shape-specific DN uptake patterns across cell lines and splenocytes and quantify in vivo DN accumulation in lymphoid organs. Together, origamiFISH-Flow offers a new tool to interrogate DN interactions with cells and tissues, while providing insights for tailoring their designs in bio-applications.

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Alexander, S., Wang, W. X., Tseng, C.-Y., Douglas, T. R., Chou, L. Y. T.. 2023-11-02. High-throughput, label-free detection of DNA origami in single-cell suspensions using origamiFISH-Flow. https://doi.org/10.1101/2023.10.31.564845

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