Expansion Microscopy with Multifunctional Polymer Dots

Abstract

Expansion microscopy (ExM) provides nanoscale resolution on conventional microscopes via physically enlarging specimens with swellable polyelectrolyte gels. However, challenges involving fluorophore degradation and dilution during sample expansion have yet to be overcome. Herein, sequential cellular targeting, gel anchoring, and high‐fidelity fluorescence reported using multifunctional polymer dots (Pdots) designed for ExM applications are demonstrated. The impressive brightness of the Pdots facilitates multicolor ExM, thereby enabling visualization of a variety of subcellular structures and neuron synapses. The average fluorescence intensities of Pdots in ExM range from ≈3 to 6 times higher than those achieved using commercially available Alexa dyes. Moreover, the fluorescence brightness and optical fluctuation are significantly improved by a surfactant‐containing expansion buffer, which enables further resolution enhancement via super‐resolution optical fluctuation imaging (SOFI). The combination of ExM and SOFI allows subcellular structures of ≈30 nm to be resolved by conventional microscopes. These results highlight the immense potential of multifunctional Pdots for ExM‐enhanced super‐resolution imaging.

Surface-Engineered Gold Nanoclusters for Stimulated Emission Depletion and Correlated Light and Electron Microscopy Imaging

“…After verifying the biocompatibility of our synthesized Au NCs (Figure S4), we investigated the surface modification capability for specific biomolecule labeling, which is important for the practical application of STED nanoprobes to bioimaging. − …”

A pH-Responsive Nanoplatform Based on Fluorescent Conjugated Polymer Dots for Imaging-Guided Multitherapeutics Delivery and Combination Cancer Therapy

“…We developed a conjugated polymer based nanoplatform with a TME-responsive CaP nanocoating to codeliver siRNAs and chemotherapeutic drug for combination cancer therapy. The conjugated polymer dots have shown excellent properties in bioapplications including super-resolution fluorescence imaging, − molecular imaging in vivo, − biosensors, − and tumor diagnosis and therapy. − In this work, the fluorescent conjugated polymer nanoparticles (CPNPs) with a high fluorescence quantum yield (∼0.60) were induced in a nanoplatform not only as cores to load siRNA but also as an indicator to track the fate of nanoplatforms in cells Scheme A illustrated the overall process for loading siRNA on surfaces of CPNPs, subsequently coating CaP shells on CPNPs surfaces to prevent therapeutic agents from leaking or degrading as they were delivered in vivo, and to obtain a controlled release of therapeutic agents (siRNA and chemotherapeutic drug HCPT) within cells.…”

Expansion microscopy with ninefold swelling (NIFS) hydrogel permits cellular ultrastructure imaging on conventional microscope

“…Another approach is through innovations in sample preparation, for example, expansion microscope (ExM) (19)(20)(21), which embeds the specimen into a hydrogel that could expand four times when dialyzed in water, realizing ~70-nm resolution. The ExM technique does not require special machinery or particular data processing, therefore providing affordable approaches for superresolution imaging in most laboratories (22)(23)(24)(25)(26)(27)(28), and has been applied in neuroscience, pathology, and mRNA discovery (29)(30)(31)(32)(33)(34)(35)(36). However, current ExM methods only manage to reach a resolution of about 70 nm, which still lags behind that in modern superresolution instruments (4,16,37,38).…”