## 1 Introduction
To date, numerous methods have been developed to isolate, detect, and characterize exosome-like vesicles. The most common protocol that is considered to be a gold standard for isolation of exosomes is differential centrifugation, which includes three main steps: low speed centrifugation during a short time period; high speed centrifugation with extended time period to remove cell debris and microvesicles; and ultra-high speed centrifugation, or ultracentrifugation (UC), at forces of 100,000 g or greater to precipitate exosomes [31; 32]. An additional centrifugation step, using sucrose or iodixanol to form a density gradient, can be coupled with differential centrifugation to further purify the sample from contaminants [33]. Size-exclusion chromatography [34], immuno-affinity capture [35], filtration [36; 37], polymer-based precipitation [38], field-flow fractionation [39], and different combinations of these methods [36; 40] have been introduced in recent years to reduce the cost and time needed for exosome isolation and achieve high yield and purity of exosome samples. Microfluidic technology is yet another new and effective technique for the non-invasive separation of exosome-like vesicles preserving the structure and composition of intact exosomes [41; 42] with the ability to process and characterize exosomes by using immunabeled entities simultaneously [43; 44; 45]. At present, the enzyme-linked immunosorbent assay (ELISA) is the gold standard for exosome quantification using their membrane proteins, such as CD63, CD9, and CD81 [46; 47; 48]. Although ELISA provides rapid analysis of exosomal membrane proteins with relatively low cost, it is difficult to capture all subpopulations of exosomes containing the protein of interest and perform fast analysis of multiple samples. To enhance efficiency in the quantification and multiplex detection of exosomes, optical methods have been developed. While nanoparticle tracking analysis (NTA) [49; 50], dynamic light scattering (DLS) [51], and flow cytometry (FC) [52; 53] can be used to obtain the size distributions and concentrations of exosomes in the fluid samples, fluorescence microscopy is used for in vitro imaging and tracking of exosomes labeled by specific fluorophores [54; 55]. Surface plasmon resonance (SPR) provides quantitative information about exosomes attached to the surface of the SPR sensor with an exosome specific antibody [56; 57]. One of the most powerful approaches to detect biomolecules with high sensitivity, specificity, and multiplexing capability is surface-enhanced Raman scattering (SERS) [58; 59; 60]. SERS is also widely used to interrogate conformational changes in molecules under various external conditions [61; 62; 63]. It is a non-invasive method, which operates in a small volume and is suitable for real-time measurements with minimum sample preparation [64; 65; 66]. SERS is based on strong amplification of Raman scattering intensity from molecules adsorbed on noble metal (Ag, Au) nanostructures or located in the vicinity of the nanostructure surface [67; 68; 69]. Specifically, huge SERS enhancement (more than \(10^{9}\)-\(10^{11}\)) can be achieved when molecules are located within an interparticle junction (\"plasmonic hot spot\") or around sharp tips [70; 71; 72]. There are two generally accepted mechanisms that contribute to SERS enhancement: (i) The electromagnetic mechanism, when local electromagnetic field is increased due to the resonance with the surface plasmons [73]; (ii) and chemical enhancement, which attributes to the charge transfer between a metal nanoparticle and molecule [74]. In addition, a molecule can be stimulated to an excited state by surface plasmons, this mechanism is assigned to molecular resonance or surface-enhanced resonance Raman scattering (SERRS) [75].