Ultrastable Fluorescent Polymer Dots for Stimulated Emission Depletion Bioimaging

Abstract

used fluorophores, such as organic dyes and fluorescent proteins. This severely limits STED nanoscopy for long-term observation of cellular events. [2] Moreover, the lateral spatial resolution of STED nanoscopy is theoretically depend on STED beam intensity, with a higher intensity resulting in a better resolution. [3] Therefore, highly photostable fluorophores are urgently desired for the wide application of STED imaging.Besides the efforts to develop more photostable dye molecules, in recent years, the application of fluorescent nanoparticles (NPs), such as quantum dots (QDs), aggregation induced emission (AIE) NPs, upconversion NPs (UCNPs) and dye hybrid plasmonic nanoparticles has been the attractive approach for STED imaging. [4] With their distinctive advantages, nanoparticles' limitations are also found for further improvements. For example, QDs and AIE NPs are bright and have enhanced antiphotobleaching properties, but the donutshaped STED beam often cause a dim "halo" by two-photon excited fluorescence, thus reducing signal to noise ratio. [4b,c] UCNPs only require a low-intensity STED beam, nonetheless, they have relatively low quantum yield and rather long lifetimes which reduced the imaging speed of STED nanoscopy. [4d] Dye hybrid plasmonic nanoparticles efficiently reduced depletion power and improved resolution, whereas heating of metal nanoparticles may lead to thermally induced NP motion and destruction of dyes and NPs. [4e,f ] Herein, we reported for the first time, the development of semiconducting polymer dots (Pdots) as a new class of ultrastable and bright imaging probe in STED nanoscopy. With the closely packed chains of hydrophobic semiconducting polymers, Pdots are emerging fluorescent NPs featuring with high brightness, good biocompatibility and fine labeling capability for a broad range of applications in chemical and biomedical sensing. [5] A wide variety of semiconducting polymers have been developed with different molecular structures, emission wavelengths and NP sizes. [6] Recently, small-sized (<13 nm) Pdots (PFO/poly(9,9-dioctylfluorenyl-2,7-diyl), PFBT/poly[(9,-9-dioctylfluorenyl-2,7-diyl)-co-(1,4-benzo-(2,1′,3)-triadiazole)], CN-PPV/poly[2-methoxy-5-(2-ethylhexyloxy)-1,4-(1-cyanovinylene-1,4-phenylene)], PFTBT5/poly[(9,9-dioctylfluorene)-co-(4,7-di-2-thienyl-2,1,3-benzothiadiazole)]) have been reported for multicolor super-resolution optical fluctuation imaging Stimulated emission depletion (STED) nanoscopy is an emerging superresolution imaging technique, which enables high spatiotemporal characterization of cellular structures and dynamics. However, its current application is limited by the lack of photostable fluorophores, which can endure strong STED illumination. Herein, a type of photobleaching-resistant semiconducting polymer dots (Pdots) for STED imaging is reported. The Pdots are synthesized by nanoprecipitation of the mixture of hydrophobic fluorescent polymer poly[{9,9-dihexyl-2,7-bis(1-cyanovinylene)fluorene}-alt-co-{2,5bis(N,N′-diphenylamino)-1,4-phenylene}] (...

Fluorescent Probes for STED Optical Nanoscopy

“…Fluorescent semiconductor polymer dots (Pdots) can also be used as STED probes thanks to their high brightness, photostability, and low cellular toxicity. Wu et al adopted Pdots for STED imaging and demonstrated high biocompatibility, photostability, and good depletability [ 110 ]. Pdots could be conjugated with molecules such as biotin and showed a very low STED power requirement (<3 mW for 70 nm resolution).…”

Designing Sub‐2 nm Organosilica Nanohybrids for Far‐Field Super‐Resolution Imaging

“…However, it remains challenging to find a class of STED nanoprobes that fulfill all these requirements in practice. For recently emerged STED nanoprobes, including quantum dots, polymer nanoparticles, aggregation‐induced emission nanoparticles, and upconversion nanocrystals, despite their distinct features, they still suffer intrinsic limitations such as large physical dimensions, high saturation intensity or low lateral resolution.…”

Near‐Infrared‐Emitting Boron‐Difluoride‐Curcuminoid‐Based Polymers Exhibiting Thermally Activated Delayed Fluorescence as Biological Imaging Probes

“…Alternatively, hydrophobic polymers exhibiting TADF can be coprecipitated with amphiphilic polymers in water, giving luminescent nanoparticles termed polymer dots (Pdots). Pdots are water‐soluble nanoparticles containing ≥50 % semiconducting polymer by weight or volume, [20] and have emerged as powerful probes for bioimaging, [20–26] biosensing, [27–30] and photodynamic therapy [31–33] . Characterized by improved photostability and high brightness, Pdots can also be functionalized for cell targeting and therapy, providing a versatile platform for theranostics [34] .…”

“…Pdots are water-soluble nanoparticles containing ! 50 %semiconducting polymer by weight or volume, [20] and have emerged as powerful probes for bioimaging, [20][21][22][23][24][25][26] biosensing, [27][28][29][30] and photodynamic therapy. [31][32][33] Characterized by improved photostability and high brightness,P dots can also be functionalized for cell targeting and therapy, providing aversatile platform for theranostics.…”