STED Nanoscopy Imaging of Cellular Lipid Droplets Employing a Superior Organic Fluorescent Probe

Abstract

Lipid droplets (LDs) are spherical organelles that participate in numerous biological processes. In order to visualize LDs on the nanoscale, nanoscopy fluorescence imaging is considered as the most attractive technique but is substantially limited by the characteristics of fluorescent probes. Thus, the development of a superior fluorescent probe that is capable of nanoscopy fluorescence imaging has attracted enormous attention. Herein, a benzodithiophene-tetraoxide-based molecule Lipi-BDTO has been developed that can easily undergo the stimulated emission depletion (STED) process and displays high photostability. These two characteristics of fluorescent probes finely satisfy the requirements of STED nanoscopy imaging. Indeed, applying the probe for STED imaging achieves a high resolution of 65 nm, belonging to one of the leading results of LDs fluorescence imaging. Furthermore, the high photostability of this fluorescent probe enables it to monitor the dynamics of LDs by time-lapse STED imaging as well as to visualize the three-dimensional (3D) spatial distribution of LDs by 3D STED imaging. Notably, the resolution of the 3D STED image represents one of the best LDs fluorescence imaging results so far. Besides STED nanoscopy imaging, the superior utility of this fluorescent probe has been also demonstrated in two-color 3D confocal imaging and four-color confocal imaging.

Fluorescent probes based on multifunctional encapsulated perylene diimide dyes for imaging of lipid droplets in live cells

“…The live-cell permeability of the SFPDIs should be closely connected with the balance between the lipophilicity of the isopropylphenyl groups and the hydrophilicity of the sulfone groups. 26…”

“…Considering the lipophilic characteristic of the perylene diimide and isopropylphenyl groups, we speculate that SFPDIs may specifically stain lipid droplets (LDs). 26 LDs, mainly comprising various neutral lipids, are cellular organelles that are essential for maintaining lipid and energy homeostasis. 27 LD-targeting fluorescent probes have been especially effective tools for the visualization of the dynamic morphology of LDs and the investigation of their related activities in real time.…”

“…The live-cell permeability of the SFPDIs should be closely connected with the balance between the lipophilicity of the isopropylphenyl groups and the hydrophilicity of the sulfone groups. 26 Considering the lipophilic characteristic of the perylene diimide and isopropylphenyl groups, we speculate that SFPDIs may specifically stain lipid droplets (LDs). 26 LDs, mainly comprising various neutral lipids, are cellular organelles that are essential for maintaining lipid and energy homeostasis.…”

“…The outstanding photostability of the SFPDIs should be closely connected with their low LUMO level and excellent anti-oxidation ability. 24,26…”

Fluorescent Probes for STED Optical Nanoscopy

“…By controlling the hydrophobicity or hydrophilicity of the probe, surface probes can be preferentially targeted to, for instance, mitochondria or nucleic acids [ 51 ]. Many specific targeting methods can thus be suggested for live cell imaging under harsh light irradiation conditions [ 52 , 53 , 54 ]. It is worth noting that FPs genetically fused to the target molecule tend to have high target specificity as well as ease of labeling.…”