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Determination of Exosome Concentration in Solution Using Surface Plasmon Resonance Spectroscopy

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Department of Applied Physics, Chalmers University of Technology, SE-412 96 Gothenburg, Sweden
Krefting Research Centre, Department of Internal Medicine and Clinical Nutrition, University of Gothenburg, SE-405 30 Gothenburg, Sweden
§ Boreskov Institute of Catalysis, Russian Academy of Sciences, 630090 Novosibirsk, Russia
Institut Curie, Centre de Recherche, CNRS, UMR168, Physico-Chimie Curie, 75005 Paris, France
Cite this: Anal. Chem. 2014, 86, 12, 5929–5936
Publication Date (Web):May 22, 2014
https://doi.org/10.1021/ac500931f
Copyright © 2014 American Chemical Society

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    Exosomes are cell-secreted nanometer-sized extracellular vesicles that have been reported to play an important role in intercellular communication. They are also considered potential diagnostic markers for various health disorders, and intense investigations are presently directed toward their use as carriers in drug-delivery and gene-therapy applications. This has generated a growing need for sensitive methods capable of accurately and specifically determining the concentration of exosomes in complex biological fluids. Here, we explore the use of label-free surface-based sensing with surface plasmon resonance (SPR) read-out to determine the concentration of exosomes in solution. Human mast cell secreted exosomes carrying the tetraspanin membrane protein CD63 were analyzed by measuring their diffusion-limited binding rate to an SPR sensor surface functionalized with anti-CD63 antibodies. The concentration of suspended exosomes was determined by first converting the SPR response into the surface-bound mass. The increase in mass uptake over time was then related to the exosome concentration in solution using a formalism describing diffusion-limited binding under controlled flow conditions. The proposed quantification method is based on a calibration and control measurements performed with proteins and synthetic lipid vesicles and takes into account (i) the influence of the broad size distribution of the exosomes on the surface coverage, (ii) the fact that their size is comparable to the ∼150 nm probing depth of SPR, and (iii) possible deformation of exosomes upon adsorption. Under those considerations, the accuracy of the concentration determination was estimated to be better than ±50% and significantly improve if the exosome deformation is negligible.

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    Mass-uptake determination using SPR; calibration of the SPR system for concentration determination; size determinations of biotinylated lipid vesicles and exosomes under different experimental conditions; concentration determination of vesicles using NTA; theoretical derivations of (i) the influence on the SPR response of the size distribution of the vesicle and exosome samples and (ii) the correction of the mass uptake estimated from the SPR signal due to surface-induced vesicle deformation; and an experimental section. This material is available free of charge via the Internet at http://pubs.acs.org.

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