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<p>Expansion microscopy (ExM) enlarges biological samples by embedding them in a swellable hydrogel, enabling nanoscale imaging of subcellular structures with standard light microscopes. This offers an accessible alternative to super-resolution methods. ExM has yet to be combined with the Oligopaint DNA fluorescence <i>in situ</i> hybridization (FISH) technology. We present an optimized ExM workflow for simultaneous Oligopaint DNA FISH and immunofluorescence (IF) in intact <i>Drosophila</i> ovaries. The protocol incorporates nucleic-acid anchoring, reliable protein retention, and digestion conditions that preserve chromatin while maintaining probe accessibility. Our approach achieves ~ 5 × expansion and strong signal retention, enabling high-resolution studies of nuclear organization.</p>

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Combining ultrastructure expansion microscopy with immunofluorescence and Oligopaint DNA FISH

  • 期刊论文

  • 发表日期：

    2026年5月21日

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  • Lorielle M. Raab,
  • Clio B. Hockens,
  • Ling Sze Lee,
  • Rose K. Runyan,
  • Emma E. Burns,
  • Nasser M. Rusan,
  • Leah F. Rosin

摘要

Expansion microscopy (ExM) enlarges biological samples by embedding them in a swellable hydrogel, enabling nanoscale imaging of subcellular structures with standard light microscopes. This offers an accessible alternative to super-resolution methods. ExM has yet to be combined with the Oligopaint DNA fluorescence in situ hybridization (FISH) technology. We present an optimized ExM workflow for simultaneous Oligopaint DNA FISH and immunofluorescence (IF) in intact Drosophila ovaries. The protocol incorporates nucleic-acid anchoring, reliable protein retention, and digestion conditions that preserve chromatin while maintaining probe accessibility. Our approach achieves ~ 5 × expansion and strong signal retention, enabling high-resolution studies of nuclear organization.