Measuring Zeta Potential of Nanoparticles: Version 1.2

This protocol describes the measurement of zeta potential for nanomaterial samples in aqueous solutions. Particle surface characteristics and charge play an important role in the particle’s physical state, stability in different media, agglomeration tendencies, and interaction with biological systems. These properties are especially important for nanomaterials used in a biological context (e.g. nanomedicines). Zeta potential measurement provides an indirect measure of the net charge and as a tool to compare batch-to-batch consistency. A particle in solution with a net charge has a layer of ions (of opposite charge) strongly bound to its surface; this is referred to as the Stern layer. A second diffuse outer layer is comprised of loosely associated ions. These two layers are collectively called the electrical double layer. As the particle moves (due to gravity, kinetic energy, and/or under an applied electrical field), there exists a boundary between the ions in the diffuse layer that move with the particle and ions that remain with the bulk dispersant. The electrostatic potential at this “slipping plane” boundary is the zeta potential. In zeta potential measurements, an electrical field is applied across the sample and the movement of the particles (electrophoretic mobility) is measured by the light scattering of the particles. The Henry equation is then used to calculate the zeta potential, : where is the electrophoretic mobility, ε is the dielectric constant, is the absolute zero-shear viscosity of the medium, is the Henry function, and κ is a measure of the ratio of the particle radius to the Debye length. This assay protocol outlines the procedure for sample preparation and determination of zeta potential. Guidelines for making successful zeta potential measurements are provided, as well as a discussion of relevant standards and data analysis. Examples of zeta potential results are illustrated for 30 nm nominal size colloidal gold and a G5 PAMAM amine-terminated dendrimer.