Towards specific quantification of exosomes using surface-based sensing

Licentiate thesis, 2014

When exosomes were first observed in 1967, they were believed to be cell debris until extensive studies demonstrated key biological functionalities related to their composition and formation mechanism. Exosomes are nanometer-sized lipid vesicles of endocytic origin, with a membrane consisting of lipids and proteins and encapsulating a cargo composed of soluble proteins and genetic material. Exosomes have been identified to play a role in intercellular communication processes such as modulating the immune system, inflammation reactions and tissue regeneration. Besides conveying new insight in fundamental biology, exosomes are also seen as promising diagnostic marker candidates. Moreover, the ability of exosomes to be taken up by a large variety of cells has directed investigations towards their use as cargo carriers in drug delivery and gene therapy applications. However, progress in these directions is hampered by the limited availability of methods capable of accurately determining exosome concentration in complex biological samples. We hereby present a method for exosome quantification using label-free surface-based sensing with surface plasmon resonance (SPR) read-out. This technique enables the determination of the concentration of analyte in solution from binding experiments operated under mass transport limited conditions. The method was evaluated and verified using synthetic lipid vesicles that served as model system for exosomes. Exosomes released from human mast cells and carrying the tetraspanin membrane protein CD63 were subsequently quantified. Additionally, liposomes and exosomes were quantified with nanoparticle tracking analysis, which is a more common method for this purpose. A main conclusion is that SPR offers a valuable complement to more established techniques, especially with respect to concentration determination of low-abundant sub-populations of exosomes present in complex biological samples.