Highly Efficient Red/NIR-Emissive Fluorescent Probe with Polarity-Sensitive Character for Visualizing Cellular Lipid Droplets and Determining Their Polarity.

Lipid droplets (LDs), which are ubiquitous organelles existing in almost all eukaryotic cells, have attracted a lot of attention in the field of cell biology over the last decade. For the biological study of LDs via fluorescence imaging, the superior LD fluorescent probes with environmental polarity-sensitive character are highly desired and powerful but are very scarce. Herein, we have newly developed such a kind of fluorescent probe named LDs-Red which enables us to visualize LDs and to further reveal their polarity information. This fluorescent probe displays the advantages of intense red/near-infrared emission, high LD staining specificity, and good photostability; thus, it would be very useful for LD fluorescence imaging application. As a result, the three-dimensional confocal imaging to visualize spatial distribution of LDs and the multicolor confocal imaging to simultaneously observe LDs and other cellular organelles have been realized using this new LD fluorescent probe. Furthermore, the polarity-sensitive emission character of this probe enables us to quantitatively determine the LD polarity via spectral scan imaging. Consequently, the cancer cells (HepG2, HeLa, and Panc02) displaying lower polarity of LDs than the normal cells (L929, U251, and HT22) have been systematically demonstrated. In addition, this polarity-sensitive probe displaying shorter fluorescence wavelengths in cancer cells than in normal cells has an important and potential ability to distinguish them.

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

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.

Super-resolution dynamic tracking of cellular lipid droplets employing with a photostable deep red fluorogenic probe.

Lipid droplets (LDs) are critical organelles involved in many physiological processes in eukaryotic cells. To visualize and study LDs, particular the small/nascent LDs, the emerging super-resolution fluorescence imaging techniques with nanoscale resolution would be much more powerful in comparison to the conventional confocal/wide-field imaging techniques. However, directly limited by the availability of advanced LDs probes, super-resolution fluorescence imaging of LDs is a practically challenging task. In this context, a superior LDs fluorescent probe named Lipi-Deep Red is newly developed for structured illumination microscopy (SIM) super-resolution imaging. This fluorescent probe features with the advantages of strong deep red/NIR emission, fluorogenic character, high LDs specificity, and outstanding photostability. These advantages enable the fluorescent probe to be finely applied in SIM super-resolution imaging, e.g. time-lapse imaging (up to 1000 frames) to monitor the LDs dynamics at nanoscale (159 nm), two-color time-lapse imaging to discover the nearby contact/interaction between LDs and mitochondria. Consequently, the fusion processes of LDs are impressively visualized at a high spatial and temporal resolution. Two kinds of contact models between LDs and mitochondria (dynamic contact and stable contact) newly proposed in the recent literatures are successfully revealed.