![dynamic light scattering protocol dynamic light scattering protocol](https://bif.wisc.edu/wp-content/uploads/sites/389/2017/11/DLS2-300x291.jpg)
The contribution of each population was extracted from the total LS intensity on the basis of the intensity weighted R h distributions. The polydispersity effect on LS data was considered. The size distributions of the particles in the samples were determined. This study focused on the viscosity of the fluid in which EVs moved during the measurement. A commercial exosome standard was used as a reference. Īssuming the validity of the above, the blood plasma and isolates from blood plasma of 3 healthy donors were analysed. The gross distribution of mass within the particles can be estimated by comparison of obtained ρ values with those of the particles with well-defined topology. Combining the DLS and SLS parameters ( R h and R g, respectively) yields the so-called shape parameter ρ (= R g/ R h). SLS, on the other side, gives another size parameter, the radius of gyration of particles ( R g). In well-defined samples (preferably monodisperse or with known polydispersity), the underlying model (based upon the movement of particles in a fluid continuum with a constant viscosity) is relevant to determine the hydrodynamic radius of the particles ( R h). The size distributions can be obtained by DLS without any a priori assumption of the particles’ chemical nature or structure. The size range of those methods (1 nm–1 µm) is consistent with the expected size of EVs. A relatively small volume of the sample (less than a milliliter) is sufficient for analysis. These techniques do not require destruction of the sample. The techniques of static (SLS) and dynamic light scattering (DLS) and their combination are commonly used in research and industry of various nano-systems. Here, the possibility to use static and dynamic light scattering (SLS and DLS, respectively) is explored to assess the size and shape of EVs in isolates as well as in blood plasma. The development of methods for minimally invasive analysis of EVs in body fluids is therefore highly desirable. This implies ethical bias and a higher cost of analysis, and thereby prevents a better understanding of these diverse biological features. Furthermore, the analytical methods for EV determination often require specific markers. Consequently, viewing EV isolates as composed of particles in the state as formed in the body is to some degree questionable. It was suggested that EVs result from self-organization of the available physico-chemical components following their exposure to various external factors. A possibility of EV transformations due to the aggressive processes of isolation from body fluids should be taken into account. The dynamics of a vesicle membrane and its contents in ex-vivo samples is poorly understood. Despite intensive investigations of these membrane particles in the last few decades, numerous issues remain unresolved, an important one being the effect of pre-analytical handling of samples on the results. The concentration of EVs in ex-vivo samples of blood plasma was estimated at 10 10 particles/mL, but may be increased as a result of various pathological conditions. It can be concluded that light scattering could be a plausible method for the assessment of EVs upon considering that EVs are a dynamic material with a transient identity.Įxtracellular vesicles (EVs) are membrane micro/nano-particles commonly found in biological samples including body fluids. This study has estimated the value of the viscosity coefficient of the medium in blood plasma to be 1.2 mPa/s. The average shape parameter of the assessed particles was found to be ρ ~ 1 (0.94–1.1 in exosome standards and 0.7–1.2 in blood plasma and EV isolates), pertaining to spherical shells (spherical vesicles). Their size and shape were assessed by using a combination of static and dynamic light scattering.
![dynamic light scattering protocol dynamic light scattering protocol](https://www.jove.com/files/ftp_upload/60257/60257fig9large.jpg)
The EVs were harvested from blood plasma by repeated centrifugation and washing of samples. The purpose of this work is to better understand the mechanisms taking place during harvesting of EVs, in particular the role of viscosity of EV suspension. Furthermore, the identity and genesis of EVs are still obscure and the relevant parameters have not yet been identified. A plausible model for the description of EV isolates has not been developed. However, the assessment methods that would yield repeatable concentrations, sizes and compositions of the harvested material are missing. Extracellular vesicles (EVs) isolated from biological samples are a promising material for use in medicine and technology.