Measure the Zeta Potential of Individual Particles with TRPS

Characterise nanomedicines, drug-delivery complexes, and bionanoparticles with unsurpassed accuracy and precision. Obtain true measurements of the size distribution, particle concentration, particle charge and charge distribution. Carry out complex analyses of heterogenous samples including characterisation of individual subpopulations without the need for laborious protocols. Perform real-time measurements of particle properties to assess subtle changes over time with high precision for quality control and assessment of product stability.  

The field of nanomedicine is progressing rapidly, and specific regulatory procedures and requirements imposed by governing bodies are likely to come into play soon. As the only technique able to provide data of sufficient quality for such purposes, TRPS is fast becoming an essential aspect in nanomedicine characterisation and quality control.
Particle zeta potential, size and concentration on a 3-axis graph.
Figure 1.
3d plot of concentration vs particle diameter and zeta potential of a trimodal mix of 150 nm and 200 nm bare polystyrene and 200 nm carboxylated polystyrene particles. The carboxylated particles carry a higher negative surface charge and hence their zeta potential is more negative than the respective zeta potentials of the bare polystyrene particles.

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Much more precise & reliable than DLS based measurement

Understand the subtle changes from different formulations.
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Simultaneous size & zeta potential measurement

Measure individual particles, on a particle-by-particle basis.
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Charge measurement in physiological-strength buffer

Measure and analyse particles in the medium that they will be used.

True zeta potential distributions provide unbiased results

In a mixture, it is common to assume that all of the particles have the same zeta potential but this is often not the case. When particles have polydisperse size and surface charge distributions, the most widely used technique (Laser doppler velocimetry, PALS) to measure the zeta potential becomes quite unreliable. The solution is to individually measure the electrophoretic mobility of a large enough number of single particles, which TRPS does very well. The electrophoretic mobility is converted to a zeta potential via their linear relationship. This results in zeta potential data that is unbiased by the size distribution and can be provided as a size vs charge plot, a concentration vs charge plot, or a 3d concentration vs charge and size. That level of information is essential for sophisticated applications like nanomedicine development and nanomedicine QA. It can be used to further identify surface biomolecular properties or biomolecular activity.
Figure 2.
Zetapotential vs particle size of bare polystyrene (CPN100), carboxylatedpolystyrene particles (CPC70, CPC100), magnetic particles (Bio-Adembeads) andmagnetic particles modified with DNA. The mix of all 5 particle types resemblesvery well the particle distributions when particle types are measuredseparately.
Figure 3.
Comparison of TRPS (top) and PALS analysis (bottom) of bimodal charged samples of 380 nm low charged bare polystyrene particles and 400 nm highly charged carboxylated polystyrene particles. There is a huge discrepancy between both techniques. Whilst TRPS can resolve the two particle types perfectly (top) with zeta-potentials agreeing well with values from unmixed samples, PALS can only measure a solution-averaged zeta-potential value (bottom).

Much more precise and reliable than PALS based zeta measurement

Phase analysis light scattering (PALS), an ensemble technique based on laser doppler velocimetry, is the best known technology for the determination of zeta potential of nanoparticle suspensions. Although readily available, ensemble techniques (e.g. PALS), as opposed to single particle measurement techniques, such as TRPS, can only measure and calculate the average particle mobility and hence detailed single particle information is lost, in particular when measuring polydisperse samples. TRPS is the only available technology that provides simultaneous in-suspension information about particle size and zeta potential on a particle-by-particle basis, guaranteeing accurate analysis of multi-modal and polydisperse samples (see figure). Whilst a mixed sample of bare and carboxylated polystyrene spheres with equivalent sizes (~400 nm) but different zeta potentials was perfectly resolved with TRPS, the two particle populations could not be distinguished with PALS.

Measure size & zeta potential simultaneously

Measuring size and zeta potential of particles simultaneously has incredible benefits for a wide range of applications and industries. TRPS is able to do this on a particle-by-particle basis with high reliability and reproducibility. It represents a new approach for understanding and researching the intrinsic behaviour of nanoscale distributions. Since these nanoparticle properties have been seen to play a central role in biological interactions, including influencing, their uptake by the target tissues/cells, TRPS allows for the investigation and better understanding of things such as nanoparticle-based delivery of microRNA. This in turn could be used for many therapeutic applications, such as overcoming drug resistance, in cancer therapy and in diagnostics. TRPS instruments are also the only devices on the market that can do this simultaneously and accurately. The high-resolution single particle size and zeta potential characterisation will positively impact developmental nanomedicine by providing a better understanding of nano-bio interactions.
Figure 4.
Zetapotential vs particle size of bare polystyrene (CPN100), carboxylatedpolystyrene particles (CPC70, CPC100), magnetic particles (Bio-Adembeads) andmagnetic particles modified with DNA. The mix of all 5 particle types resemblesvery well the particle distributions when particle types are measuredseparately.
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Figure 5.
Simultaneous TRPS size and zeta potentialmeasurements of a pentamodal mix of bare (CPN) and carboxylated (CPC)polystyrene particles in PBS (phosphate buffered saline).

Charge measurement in physiological-strength buffer

Particle dispersions and formulations are stabilized by electrostatic repulsion, steric hindrance, or a combination of these two forces. Particles will eventually aggregate in the absence of sufficient stabilisation. Researchers use zeta potential as an indicator of electrostatic stabilisation of particles. Zeta potential is a modelled quantity derived by measuring electrophoretic mobility of particles in suspension. Electrophoretic mobility is critically dependent on particle and solution properties (ionic strength, ionic composition, and viscosity). Thus, it’s important to perform zeta analysis for nanoparticles or nano-formulations in physiological buffer(s) designed for biological applications.

High-resolution single particle zeta analysis will provide an advanced understanding of particle behaviour in different pH and salt conditions and aid in monitoring particle corona evolution over time for biokinetic studies. Moreover, determination of accurate charge on each particle could provide potential insights into nano-bio interactions in varying physiological buffers, critical for determining particle stability and uptake; impacting the overall delivered particle dosage.

Fine scale precision

TRPS alone provides the means to distinguish particles through fine scale mapping of zeta potential of individual particles. The surface charge of membranous vesicles arise from the combined net charge of extrinsic moieties on the vesicle surface. Using TRPS, subtle changes in surface charge can be resolved as the membrane composition is altered.  

TRPS can also be used to monitor changes in zeta potential through particle-ligand interaction in real time. This is demonstrated below in the binding of biotinylated single stranded DNA (35 bases) to the surface of a streptavidin coated particle. The diffusion dependent binding reaction goes to completion within approximately 170 sec after the addition of DNA at 30 sec, the binding of the DNA coincides with a reduction in the zeta potential. The resolution limit was 10% of the oligo coverage. For example, the resolution limit was 44 oligos per particle for particles with a DNA/particle ratio of 400. The control reaction which uses streptavidin that has been enzymatically inactivated by proteinase K shows no DNA interaction. With longer oligos the sensitivity is increased. TRPS provides researchers with the tool for advanced charge studies of particle-particle interactions, zeta based vesicle characterisation, receptor-ligand interactions that may result in altered charge of the carrier particle, charge reporting antibody-antigen reactions and aptamer based detection technologies.
Figure 6.
Kinetics of biotinylated DNA reaction onto magnetic Bio-Adembeads was monitored via particle zeta potential measurements and compared with a control where streptavidin on the surface of the Bio-Adembeads was enzymatically cleaved before DNA reaction. Approximately 100 sec after DNA addition to the fluid cell the moving average of the zeta potential (averaged over 25 particles) levelled at approximately -20 mV, indicating reaction saturation. The zeta potential for the control does not change with time.
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Introducing the Exoid

The Exoid is the next generation of TRPS device. The Exoid has the proven quality TRPS technology developed with the previous generation, qNano, but significantly improves the user experience making TRPS measurement easier than ever before.
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