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Versatile Microfluidics Platform for Enhanced Multitarget Super-Resolution Microscopy
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DOI：10.1021/acsnano.5c18697.png)
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DNA-based Point Accumulation for Imaging in Nanoscale Topography (DNA-PAINT) is a powerful variant of single-molecule localization microscopy (SMLM) that overcomes the limitations of photobleaching, offers flexible fluorophore selection, and enables fine control of imaging parameters through tunable on- and off-binding kinetics. Its most distinctive feature is its capacity for multiplexing, typically implemented through sequential imaging of targets using an Exchange-PAINT. This technique involves assigning orthogonal DNA strands to different targets within a sample and then sequentially adding and removing complementary imager strands that are specific to only one target at a time. However, manual Exchange-PAINT workflows are often inefficient, prone to drift and variability, and lack reproducibility. Here, we introduce a custom compressed-air-driven microfluidics system specifically designed for multiplexed SMLM. Featuring a stackable and modular design that is, in principle, not limited by the number of channels, the system ensures robust, reproducible, and material-efficient buffer exchange with minimal dead volume. It operates in both manual and automated modes and can be readily adapted to a wide range of commercial and custom microscopes, including wide-field, confocal, STED, MINFLUX and other platforms. We demonstrate robust 5-plex Exchange-PAINT imaging in cancerous U2OS cells, and importantly, we establish multiplexed nanoscale imaging in fragile primary cardiomyocytes. These applications demonstrate that the platform enables reliable super-resolution multiplexing in physiologically relevant systems and supports detailed nanoscale analysis in complex primary cells.
Keyword:
super-resolution microscopy
DNA-PAINT
single-moleculelocalization microscopy
multiplexed imaging
cardiomyocytes
microfluidics
SMLM automation
期刊
IF:
16
论文数:
2.6W
被引数:
25.6W
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