Comparing Concentration Measurements between the NanoSight NS300 and Zetasizer Ultra

As a Field Application Scientist, I am often asked some form of the following question: Between NanoSight NS300 and Zetasizer Ultra, what is the “better” technology to measure particle concentration? My answer to this question is always the same: It depends.

This is an understandable question, because at first glance, it may not be clear what the difference is between the two instruments. Both instruments use light scattering to measure particles in a liquid, and both use Brownian motion to determine hydrodynamic size. However, the particle concentration measurement principles between the two technologies are fundamentally different.

NTA vs MADLS

To measure particle concentration, NanoSight uses Nanoparticle Tracking Analysis (NTA) to directly count particles in a known three-dimensional scattering volume, whereas the Zetasizer uses Multi Angle Dynamic Light Scattering (MADLS) to count photons, and calculates particle concentration from other measured or known sample properties, including size, refractive index, viscosity, and buffer scatter rate.

Size, Polydispersity, and Viscosity

From these two approaches, both instruments measure particle concentration in particles per milliliter (particles/mL). However, the differences between the two technologies lend themselves to different ideal sample types, mainly relating to size and polydispersity.

NanoSight excels at measuring particle concentration for particles within the ideal size and concentration range, which is refractive index-dependent, but is approximately 50-500 nm for size and 108-109 particles/mL for concentration. NanoSight is better able to count particles in polydisperse samples, because concentration is determined directly from particle counts, and both dim and bright particles are counted equally.

In contrast, polydispersity has a larger influence on the Zetasizer Ultra particle concentration measurement, because concentration is calculated from the measured size, and polydispersity can influence size accuracy. However, the lower sizing limit of detection for the Zetasizer Ultra is much lower than NanoSight and can measure sub-50 nm particles with ease.

Sample viscosity is also an important property for the Zetasizer Ultra particle concentration measurement. Although viscosity is used in both technologies to measure size, it only affects particle concentration on the Zetasizer Ultra because concentration is calculated from size, whereas the NanoSight does not use particle size to determine particle concentration.

Ideal Samples

Ideal sample types for NanoSight include monodisperse to polydisperse exosomes, sub-micron-sized protein aggregates, in-process liposomes, nanobubbles and larger viral vectors like Lentivirus and Adenovirus. Ideal sample types for the Zetasizer Ultra are smaller viral vectors like Adeno Associated virus (AAV) and Virus-Like Particles (VLPs), and any other type of small, monodisperse nanoparticle.

For many common applications, such as purified liposomes, polystyrene latex, and other monodisperse nanoparticles, both instruments are likely to excel and produce identical particle concentration results. The ideal instrument to track nanoparticle concentration may change with the stage of drug development or purification level, as sample properties vary from early research, to in-process development, all the way to the final drug product. For cases like these, it is valuable to have both the NanoSight and Zetasizer operating in the same lab.

Further reading

  1. Particle concentration measurements on the Zetasizer Ultra – How it works
  2. Measuring the concentration of Adeno-Associated Virus (AAV) with multi-angle dynamic light scattering (MADLS)
  3. ISO19430 – Particle Tracking Analysis Method