Plasmon-Enhanced Expansion Microscopy

Expansionmicroscopy (ExM) is a rapidly emerging super-resolutionmicroscopy technique that involves isotropic expansion of biologicalsamples to improve spatial resolution. However, fluorescence signaldilution due to volumetric expansion is a hindrance to the widespreadapplication of ExM. Here, we introduce plasmon-enhanced expansionmicroscopy (p-ExM) by harnessing an ultrabright fluorescent nanoconstruct,called plasmonic-fluor (PF), as a nanolabel. The unique structureof PFs renders nearly 15000-fold brighter fluorescence signal intensityand higher fluorescence retention following the ExM protocol (nearly76%) compared to their conventional counterparts (<16% for IR-650).Individual PFs can be easily imaged using conventional fluorescencemicroscopes, making them excellent digital labelsfor ExM. We demonstrate that p-ExM enables improved tracing and decryptingof neural networks labeled with PFs, as evidenced by improved quantificationof morphological markers (nearly a 2.5-fold increase in number ofneurite terminal points). Overall, p-ExM complements the existingExM techniques for probing structure-function relationshipsof various biological systems.

Automated summary

Expansion microscopy (ExM) is a super-resolution technique that expands biological samples to improve spatial resolution. However, fluorescence signal dilution is a challenge for ExM. In this study, plasmon-enhanced expansion microscopy (p-ExM) is introduced using an ultrabright fluorescent nanoconstruct called plasmonic-fluor (PF). PFs have a unique structure that allows for significantly brighter fluorescence signal and higher retention compared to conventional counterparts. p-ExM enables improved tracing and decryption of neural networks labeled with PFs for better analysis of structure-function relationships in biological systems.