Microfluidic Plasmonic Bio-Sensing of Exosomes by Using a Gold Nano-Island Platform
A bio-sensing method, based on the plasmonic property of gold nano-islands, has been developed for detection of exosomes in a clinical setting. The position of the gold plasmon band in the UV-Visible spectrum depends on the size and shape of gold nanoparticles as well as on the surrounding environment. By adsorbing various chemical entities, or binding them, the gold plasmon band will shift toward longer wavelengths and the shift is proportional to the concentration. Exosomes transport cargoes of molecules and genetic materials to proximal and distal cells. Presently, the standard method for their isolation and quantification from body fluids is by ultracentrifugation, not a practical method to be implemented in a clinical setting. Thus, a versatile and cutting-edge platform is required to selectively detect and isolate exosomes for further analysis at clinical level. The new sensing protocol, instead of antibodies, makes use of a specially synthesized polypeptide (Vn96), to capture and quantify the exosomes from different media, by binding the heat shock proteins from exosomes. The protocol has been established and optimized by using a glass substrate, in order to facilitate the next stage, namely the transfer of the protocol to a microfluidic environment. After each step of the protocol, the UV-Vis spectrum was recorded and the position of gold Localized Surface Plasmon Resonance (LSPR) band was measured. The sensing process was modelled, taking into account the characteristics of the nano-island structure, prepared by thermal convection and annealing. The optimal molar ratios of the most important chemical entities, involved in the detection of exosomes were calculated as well. Indeed, it was found that the results of the sensing process depend on the two major steps: the molar ratios of streptavidin to biotin-PEG-Vn96 and, the final step, the capture of exosomes by the biotin-PEG-Vn96 complex. The microfluidic device designed for sensing of exosomes consists of a glass substrate, sealed by a PDMS layer that contains the channel and a collecting chamber. In the device, the solutions of linker, cross-linker, etc., are pumped over the gold nano-islands and an Ocean Optics spectrometer is used to measure the position of the Au plasmon band at each step of the sensing. The experiments have shown that the shift of the Au LSPR band is proportional to the concentration of exosomes and, thereby, exosomes can be accurately quantified. An important advantage of the method is the ability to discriminate between exosomes having different origins.
Digital Object Identifier (DOI): doi.org/10.5281/zenodo.1316742Procedia APA BibTeX Chicago EndNote Harvard JSON MLA RIS XML ISO 690 PDF Downloads 439
 C. Thery, S. Amigorena, G. Raposo, A. Clayton, “Isolation and Characterization of Exosomes from Cell Culture Supernatants and Biological Fluids,” Curr. Protoc. Cell Biology, Chapter 3: Unit 3, 22, 2006.
 E. Van der Pol, A. N. Boing, P. Harrison, A. Sturk, R. Nieuwland, “Classification, Functions, and Clinical Relevance of Extracellular Vesicles,” Pharmacol. Rev., 64, pp. 676–705, 2012.
 S. E. L. Andaloussi, I. Mager, X. O. Breakefield, M. J. Wood, “Extracellular vesicles: biology and emerging therapeutic opportunities” Nat. Rev. Drug Discov., 12, pp. 347–57, 2013.
 A. Ghosh, M. Davey, I. C. Chute, S. G. Griffiths, S. Lewis, “Rapid Isolation of Extracellular Vesicles from Cell Culture and Biological Fluids Using a Synthetic Peptide with Specific Affinity for Heat Shock Proteins” PLoS ONE, vol. 9(10): e110443,2014.
 N. Nath, A. Chilkoti, “Label-Free Biosensing by Surface Plasmon Resonance of Nanoparticles on Glass: Optimization of Nanoparticle Size,” Anal. Chem., 76(18), pp. 5370–5378, 2004.
 N. Nath, A. Chilkoti, “Label Free Colorimetric Biosensing Using Nanoparticles,” J. Fluoresc., 14(4), pp. 377–389, 2004.
 F.Frederix, J.M.Friedt, K.H.Choi, W.Laureyn, A.Campitelli, D.Mondelaers, G.Maes, G. Borghs, “Biosensing Based on Light Absorption of Nanoscaled Gold and Silver Particles,” Anal. Chem., 75 (24), pp.6894–6900, 2003.
 J. C. Hulteen, D. A. Treichel, M.T. Smith, M.L. Duval, T.R. Jensen, R. P. Van Duyne, “Nanosphere Lithography: Size-Tunable Silver Nanoparticle and Surface Cluster Arrays,” J. Phys. Chem. B, 103 (19), pp.3854–3863,1999.
 M.D. Malinsky, K.L. Kelly, G.C. Schatz, R.P. Van Duyne, “Chain Length Dependence and Sensing Capabilities of the Localized Surface Plasmon Resonance of Silver Nanoparticles Chemically Modified with Alkanethiol Self-Assembled Monolayers,” J. Am. Chem. Soc, 123 (7), pp.1471–1482,2001.
 B. Sepulveda, P.C. Angelome, L.M. Lechuga, L.M. Liz-Marzan, “LSPR-based nanobiosensors,” Nano Today, 4, pp. 244-251, 2009.
 K.M.Byun, “Development of nanostructured plasmonic substrates for enhanced optical biosensing,” J. Opt. Soc. of Korea, 14, pp. 65-76, 2010.
 S. Unser, I. Bruzas, J. He, L. Sagle, “Localized Surface Plasmon Resonance biosensing: Current challenges and approaches,” Sensors, 15, pp. 15684-15716, 2015.
 J. Ozhikandathil, S. Badilescu, M. Packirisamy, “Gold nanoisland structures integrated in a lab-on-a-chip for plasmonic detection of bovine growth hormone, J. of Biomedical Optics, 17(7), 077001, 2012.
 R. Duraichelvan, B. Srinivas, S. Badilescu, R.J. Ouellette, A. Ghosh, M. Packirisamy, “ Exosomes Detection by a Label-free Localized Surface Plasmonic Resonance Method”, ECS Trans., 75(17), pp.11-17, 2016.