Quantification of particles is of interest in many applications, such as viruses and virus-like particles for gene delivery and biological nanoparticles or inorganic particles for drug delivery and imaging. Particle concentration in batch mode can be a high-throughput tool for screening before investing in method development for absolute concentration from size-exclusion chromatography combined with multi-angle light scattering (SEC-MALS).
Particle concentration is now calculated by DYNAMICS® 7.9 from the hydrodynamic radius, measured by dynamic light scattering (DLS), and the total static light scattering (SLS) intensity which can be determined with the DynaPro® Plate Reader III or NanoStar®. DYNAMICS uses the Lorenz-Mie model for a solid sphere for determining the particle concentration based on the hydrodynamic radius and SLS intensity at the instrument’s scattering angle.
The parameters required to perform this calculation are stored in the DYNAMICS Sample definition. In the Samples parameter node, select “Yes” to enable the Particle Concentration Calculation for a particular Sample. Then, enter the Particle Material and Shape from the choices provided. Since DYNAMICS uses the absolute refractive index (aRI) of the Sample and its corresponding Solvent in the calculations, this value must be specified for each sample. The dn/dc, which is required for Mw-S calculations in DYNAMICS, is not used for particle concentration.
Input Parameters to
DYNAMICS provides a library of materials from which to choose for each Sample definition. Materials include protein, lipid vesicles, DNA/RNA, extracellular vesicle, polystyrene, and gold. Users may also specify their own values by selecting “Other” as the material. The aRI of viruses and virus-like particles can vary depending on the payload. When measuring adeno-associated virus (AAV), for example, we recommend specifying an aRI of 1.43 for empty AAV and 1.48 for AAV filled with DNA payload.
To display the particle concentration of your sample, right-click on the data grid table and choose the Table Settings for Particle Concentration. Particle concentration is displayed in a column in the data grid for each measurement in units of 1/mL.
The graph shows results for particle concentration calculations for filled and empty adeno-associated virus (AAV) using SEC-MALS, a NanoStar, and a DynaPro Plate Reader III.
Particle concentrations for AAV calculated using the Plate Reader and NanoStar match well with those from the absolute method of SEC-MALS, allowing DLS to be used for high-throughput screens for aggregate content with no need for method development.