We have developed a novel nanoparticle-tracking-analysis (NTA) technology to visualize and analyze—in terms of both size and scattered intensity—nanoscale particles in liquid with little preparation and at low cost. NTA allows analysis of a wide range of particle types in a wide variety of solvents. The only user input required are the sample temperature and solvent viscosity. (Multi-angle measurements are superfluous.)
Our technique is based on laser-light-scattering microscopy where instruments can visualize and dynamically size individual particles in the range of 10–1000nm (dependent on material). The Brownian motion of every particle (which appears as point scatterers) is tracked separately but simultaneously using a CCD camera, from which a high-resolution plot of the particle-size distribution (and profile changes in time during, for instance, aggregation or dissolution) is obtained.1 Sample pretreatment is minimal, requiring only dilution with a suitable solvent to an acceptable concentration (between 108 and 109 particles/ml, depending on sample type). All particle types can be measured provided that they scatter sufficient light to be visible (i.e., they must not be index-matched).2
The new NTA technique is centered on a sample-analysis module (see Figure 1) in which a laser diode is configured to pass light through a 250μl liquid sample containing the nanoparticles. Particles in the beam are visualized by a conventional optical microscope aligned normally to the beam axis. This setup collects light scattered from every particle in the field of view as they move rapidly under Brownian motion: see Figure 2(a).
Figure 1. Laser module showing the beam passing through a sample.
Figure 2. (a) Nanoparticle suspension in the path of the laser beam captured by a microscope, (b) trajectories of individual-particle Brownian motion plotted by the tracking-analysis program, and (c) size-distribution profile based on particle trajectories.
The typically 20–60s video3 (30 frames/s) is analyzed on a frame-by-frame basis (see video4) using a proprietary analysis program in which each particle is identified and located automatically and its movement tracked: see Figure 2(b). The mean squared displacement for each particle can thus be determined. The diffusion coefficient (Dt) and the sphere-equivalent hydrodynamic radius (rh) can be determined using the Stokes–Einstein equation,
where KB is Boltzmann's constant, T the temperature, and η the viscosity. Figure 2(c) shows the resulting particle-size distribution.
By plotting particle size against scattered intensity, particle mixtures can be resolved to a size ratio of better than 1.3. For example, a mixture of 100 and 200nm particles can be shown—following analysis of many hundreds of particles over a 30s period—as a 3D plot with the number concentration along the vertical axis (see Figure 3).
Figure 3. Particle-size-distribution profile of a mixture of 100 and 200nm particles (referred to as 100's and 200's, respectively) analyzed over a 30s period as a function of particle intensity, allowing dimer detection.
The NTA particle-by-particle approach avoids the intensity-weighted averaging assumptions of photon-correlation spectroscopy (also known as dynamic-light scattering) and provides a unique image, going beyond existing methods in assessing polydisperse systems. The technique uniquely allows the user a simple and direct qualitative view of the sample (perhaps to validate data obtained from other techniques), from which an independent and immediate quantitative estimation of sample size, size distribution, and concentration can be obtained.
Sample types that have been measured by NTA include pigments in inks and paints (e.g., titanium dioxide), ferritin molecules, metal oxides (in magnetic-storage media), precursor chemicals for wafer fabrication, multi-walled carbon nanotubes, fuel additives (zinc dioxide), cosmetics and healthcare products, foodstuffs (microemulsions), ceramics, quantum dots, and polymers and colloids of different types. Many of these materials were suspended in nonaqueous solvents, such as toluene and heptanes. Most recently, the technique was used successfully to characterize nanoemulsions and drug-delivery particles, detect and count viruses in vaccine production and gene-therapy research, investigate the formation of nanoparticles using laser-ablation techniques, and analyze nanoparticles in nanoparticle-toxicity studies.
Further development work is aimed at providing simultaneous measurement of nanoparticle electrophoretic mobility and detection of fluorescent nanoparticles.5
Bob Carr has a PhD in microbial biochemistry and is founder of NanoSight. He has 30 years of experience in the detection and analysis of particles and biosensor research, having worked for a leading government research establishment in the biodetection sector as well on a range of industrial-contract research projects and collaborations.
5. Bob Carr, Patrick Hole, Andrew Malloy, Jonathan Smith, Andrew Weld, Jeremy Warren, The real-time, simultaneous analysis of nanoparticle size, zeta potential, count, asymmetry and fluorescence in liquids, Part. Syst. Anal. 2008. In press