Nanoparticle Measurement Through Visualization

The analysis of nanoparticles is a ubiquitous requirement in a broad range of industry sectors. Product performance and stability frequently depend on the ability to manufacture particle suspensions to fine tolerances without the presence of contaminants or aggregates. Foremost in such analyses are particle size and size distribution measurement for which a number of techniques are well-established and commonly employed in routine quality control as well as in a research and development environment.

The two most common methods are photon correlation spectroscopy (PCS), also referred to as dynamic light scattering (DLS), and electron microscopy (EM), usually scanning electron microscopy (SEM).

PCS examines the light scattered from particles, but rather than giving the particle size on a particle-by-particle basis, it produces an average figure. It does not provide a visualization of the particles and simply detects the rate of change of interference resulting from particle Brownian motion. It is a very strong technique for looking at samples with a very narrow range of particle sizes (monodispersed), but has problems with samples with a range of sizes (polydispersed) where the large particles dominate the average produced and hence bias the technique toward a few larger or contaminant particles. EM is a very exact method of measuring dry particles, but often may require significant sample preparation prior to inspection and measurement.

Improved technique

Nanoparticle tracking analysis (NTA, NanoSight Ltd., Wiltshire, U.K.) is a method of visualizing and analyzing particles in liquids that relates the rate of Brownian motion to particle size. The rate of movement is related only to the viscosity of the liquid, the temperature, and the size of the particles. It is not influenced by particle density. Compared to other light scattering techniques, like PCS, higher- resolution particle size distribution profiles can be obtained.

It is often claimed that a new instrumental technique is the easiest to use, producing the best results in the shortest amount of time, but in the case of NTA, this really is the case. NTA enables the study of individual particles from 10 to 1000 nm rather than the averaging techniques employed by PCS, also known as DLS. Analysis of 10-nm particles is only possible for particles with a high refractive index such as gold and silver, while the upper size limit is restricted by the limited Brownian motion. At 1 μm, particles move very slowly; hence the accuracy of the technique starts to diminish. The viscosity of the solvent also plays a role in determining the upper size limit since it, too, influences the movement of the particles.

Populations of particles are studied rather than conducting a search for one or two contaminant particles in a large volume. In the ideal measurement, a concentration of 106 to 109 particles per milliliter is used. This equates to less than 1 wt%, which means that, for some applications, the sample needs to be diluted. Care should be taken when diluting a sample since this may sometimes lead to particle aggregation.

Sample pretreatment is minimal; dilution with a suitable solvent to an acceptable concentration range of between 105 and 1010 per milliliter is required. Accurate and reproducible analyses can be obtained from video images of only a few seconds’ duration, and the results allow particle number concentration to be recovered. Given the close to real-time nature of the technique, particle– particle interactions are accessible, as is information about sample aggregation and dissemination. All particle types can be measured and in any solvent type, provided the particles scatter sufficient light to be visible (i.e., are not index matched).

The technique is robust, low cost, and an attractive alternative or complement to some of the higher-cost and more complex methods of nanoparticle analysis, such as photon correlation spectroscopy and electron microscopy. The technology provides the user with a simple and direct qualitative view of the sample under analysis (perhaps to validate data obtained from other techniques such as PCS) and from which an independent quantitative estimation of sample size, size distribution, and concentration can be immediately obtained.

Figure 1 - NTA flow cell.

Required hardware

NTA employs a specially designed flow cell that is mounted on a conventional optical microscope equipped with a charge-coupled device (CCD) camera capable of operating at 30 frames per second (see Figure 1). A suitably prepared sample of nanoparticles in solution is injected into the cell. The beam from a laser diode is passed through the liquid sample cell (Figure 2). Particles in the beam produce light scattering, which is viewed with a suitable optical microscope and CCD camera against a proprietary metallized surface (Figure 3).

Figure 2 - Loading a sample into the flow cell.

Figure 3 - Schematic of operation of the NTA system.

The nanoparticles move in the beam of the laser. They move randomly under solvent bombardment (Brownian motion) at a speed related to their size. The intensity of light scattered by a nanoparticle is related (through power law) to its size. Small particles will move faster and farther than large particles. The distance moved by each particle is measured and the average is determined. The particle diffusion coefficient is calculated. This relates to a sphere equivalent to the hydrodynamic diameter of the Stokes-Einstein equation.

Nanoparticles are too small to be imaged by the microscope. They are seen as point scatterers moving under Brownian motion. Larger particles will scatter significantly more light than smaller particles, while the speed of the particles varies significantly with particle size.

Figure 4 - Video capture illustrating Brownian motion.

Figure 5 - Trajectory of each individual particle as derived from the series of CCD images captured over a few seconds.

Figure 6 - Final particle size distribution, which is calculated from the sum of the individual measurements.

The process of collection through analysis is illustrated in Figures 4–6. Figure 4 shows the video capture illustrating the Brownian motion. Figure 5 shows the trajectory of each individual particle as derived from the series of CCD images captured over a few seconds. The final particle size distribution, which is made up from all the separate measurements, is shown in Figure 6.

The scales shown are linear. The horizontal scale runs from 1 to 1000 nm, while the vertical scale is a direct number count of particles, each derived from an individual measurement.