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<p>Three-dimensional (3D) nanoscale imaging reveals the detailed morphology of subcellular structures; however, conventional single-molecule localization microscopy is constrained by limited axial resolution. Here we introduce ROSE-3D, an interferometric localization approach that enables isotropic 3D super-resolution imaging with uniform performance across the entire depth of field. Compared with conventional astigmatism-based methods, ROSE-3D improves lateral localization precision by 2–6 times and axial precision by 3.5–8 times over a depth of field of approximately 1.2 μm. Leveraging its multicolor and whole-cell imaging capabilities, ROSE-3D resolves, in situ, the nanoscale organization of nuclear lamins and the assemblies of mitochondrial fission-related protein DRP1. These results establish ROSE-3D as a powerful tool for interrogating nanoscale cellular architecture.</p>

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Molecular-scale isotropic 3D super-resolution microscopy via interference localization

  • 期刊论文

  • 发表日期：

    2025年12月2日

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  • Shihang Luo,
  • Xian’ao Zhao,
  • Yuanyuan Li,
  • Chunyan Fan,
  • Ruina Liu,
  • Ran Gong,
  • Weixing Li,
  • Nana Ma,
  • Zhenghong Yang,
  • Tao Xu,
  • Wei Ji,
  • Lusheng Gu

摘要

Three-dimensional (3D) nanoscale imaging reveals the detailed morphology of subcellular structures; however, conventional single-molecule localization microscopy is constrained by limited axial resolution. Here we introduce ROSE-3D, an interferometric localization approach that enables isotropic 3D super-resolution imaging with uniform performance across the entire depth of field. Compared with conventional astigmatism-based methods, ROSE-3D improves lateral localization precision by 2–6 times and axial precision by 3.5–8 times over a depth of field of approximately 1.2 μm. Leveraging its multicolor and whole-cell imaging capabilities, ROSE-3D resolves, in situ, the nanoscale organization of nuclear lamins and the assemblies of mitochondrial fission-related protein DRP1. These results establish ROSE-3D as a powerful tool for interrogating nanoscale cellular architecture.