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Multiplexed and millimeter-scale superresolution imaging of cells and tissue sections via prism-illumination and microfluidics-enhanced DNA-PAINT

Superresolution microscopy (SRM) has become an enabling tool for biomedical research. A major limitation of SRM, however, is the small field-of-view (FOV), typically ∼50µm x 50µm and up to ∼200µm x 200µm in recent attempts, hampering its use in imaging large cell populations or clinical tissues. Here we report PRism-Illumination and Microfluidics-Enhanced DNA-PAINT (PRIME-PAINT) for efficient, multiplexed SRM across millimeter-scale FOVs. Unlike existing SRM, PRIME-PAINT uses prism-type illumination for robust DNA-PAINT with single FOVs up to a half millimeter. Through stitching, imaging over mm2 FOVs can be completed in as little as an hour per target. The on-stage microfluidics not only facilitates multiplexing but enhances image quality, particularly for tissue sections. We demonstrate the utility of PRIME-PAINT by analyzing ∼106 caveolae structures in ∼1,000 cells and imaging entire pancreatic cancer lesions from patient tissue biopsies. Thus, we expect PRIME-PAINT to be useful toward building multiscale, Google-Earth-like views of biological systems.