Since their discovery, extracellular vesicles have gained considerable scientific interest as a novel drug delivery system. In particular, outer membrane vesicles (OMVs) play a critical role in bacteria-bacteria communication and bacteria-host interactions by trafficking cell signalling biochemicals (i.e. DNA, RNA, proteins). Although previous studies have focused on the use of OMVs as vaccines, little work has been done on loading them with functional nanomaterials for drug delivery. We have developed a novel drug delivery system by loading OMVs with gold nanoparticles (AuNPs). AuNPs are versatile nanoparticles that have been extensively used in disease therapeutics. The particles were loaded into the vesicles via electroporation, which uses an electric pulse to create a short-lived electric field. The resulting capacitance on the membrane generates pores in the lipid bilayer of the OMVs allowing AuNPs (or any nanoparticle under 10 nm) inside the vesicles. Closure of the pores of the lipid membrane of the OMVs entraps the nanoparticles as cargo. Transmission electron microscopy was used to confirm the loading of AuNPs inside the OMVs and dynamic light scattering (DLS) and cryogenic scanning electron microscopy (cryo-SEM) verified the size and integrity of the OMVs. This is the first report to load nanoparticles into OMVs, demonstrating a potential method for drug delivery.

Quantum dots are attractive alternatives to organic fluorophores for the purposes of fluorescent labeling and the detection of biomarkers. They can also be made to specifically target a protein of interest by conjugating biomolecules, such as antibodies. However, the majority of the fluorescent labeling using quantum dots is done using toxic materials such as cadmium or lead due to the well-established synthetic processes for these quantum dots. Here, we demonstrate the use of indium phosphide quantum dots with a zinc sulfide shell for the purposes of labeling and the detection of exosomes derived from the THP-1 cell line (monocyte cell line). Exosomes are nano-sized vesicles that have the potential to be used as biomarkers due to their involvement in complex cell processes. However, the lack of standardized methodology around the detection and analysis of exosomes has made it difficult to detect these membrane-containing vesicles. We targeted a protein that is known to exist on the surface of the exosomes (CD63) using a CD63 antibody. The antibody was conjugated to the quantum dots that were first made water-soluble using a ligand-exchange method. The conjugation was done using carbodiimide coupling, and was confirmed using a range of different methods such as dynamic light scattering, surface plasmon resonance, fluorescent microscopy, and Fourier transform infrared spectroscopy. The conjugation of the quantum dot antibody to the exosomes was further confirmed using similar methods. This demonstrates the potential for the use of a non-toxic conjugate to target nano-sized biomarkers that could be further used for the detection of different diseases.

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