Imaging and Intracellular Tracking of Cancer Derived Exosomes Using Single Molecule Localization Based Super Resolution Microscope

Exosomes are small membrane vesicles secreted by cells and enriched with plenty of proteins. Considering their significant roles in different physical activities and potential value for diagnostic drug delivery, researchers have put great efforts in in-vitro tracking and contents analysis of exosomes. Recently, the emergence of different kinds of super-resolution microscopy provides powerful tools for exosome study.

Here, researchers from Southeast University, China demonstrate the application of single molecule localization based super resolution imaging technique (PALM/STORM) in the imaging and tracking of cancer derived exosomes. In the experiment, first, cancer-derived exosomes are extracted from the culture media of tumor cells. Then the exosome membrane receptors are labeled with photoswitchable probes, which allow super resolution imaging of these membrane receptors via photoactivated localization microscopy (PALM) or stochastic optical reconstruction microscopy (STORM). By using human breast cancer cell derived exosomes, the researchers demonstrated simultaneous dual color PALM/STORM imaging of two kinds of membrane receptors on the exosome membrane. Moreover, the successful labeling and imaging of exosomes make it possible to observe the interaction between cancer derived exosomes and normal cells. Meanwhile, they realized the colocalization of cancer derived eosomes and lysosmes in recipent cells with PALM/STORM imaging. Since exosomes play a vital role in intercellular communications, they anticipate that the presented PALM/STORM based imaging and tracking of exosomes holds a great potential in the investigation of the mechanism of exosome mediated cancer metastasis.

TIRF and PALM/STORM imaging of HeLa exosomes

(a) Schematic illustration of indirect IF labeling of CD63 and exosome membrane stained with CM-Dil. (b) TIRF image of HeLa-derived exosomes stained with CM-Dil. (c) IF labeling of CD63 with Alexa Fluor 647, conventional diffraction-limited image (upper right) and PALM/STORM image (low left). (d) Immunofluorescence control sample without primary antibody for specificity detection, CM-Dil channel. (e) Immunofluorescence control sample without primary antibody for specificity detection, Alexa Fluor 647 channel. (f) Wide field TIRF image of CD63 on a single exosome, (g) PALM image of the same region in (f). (h) Cross-sectional profiles of the exosome shown in f and g.