DLS and Zeta potential method development with practical examples

• Benjamin Lynch - Application Scientist, Malvern Panalytical

Method development for DLS and ELS: Insights from our Zetasizer Masterclass

Particle size and Zeta potential are essential parameters for pharmaceutical manufacturers to control. Particle size can have a huge impact on bioavailability, and Zeta potential analysis helps you optimize drug delivery by controlling particle interactions and ensuring uniform dispersion. Dynamic light scattering (DLS) and electrophoretic light scattering (ELS) are reliable tools to complete these measurements – if you get your methods right. Get your method development wrong, however, and you could be confronted by:

• Inaccuracies in your results
• Inconsistencies between measurements
• Increased noise in your data
• Non-compliance with regulatory requirements

In our recent Zetasizer Masterclass, our Application Scientist, Benjamin Lynch, discussed how the Zetasizer Advance range helps you avoid these pitfalls, and the factors to consider when developing a robust, reliable method for DLS or ELS analysis.

Key takeaways

• Dynamic light scattering (DLS) measures the size of particles in a sample by passing a laser beam through it and measuring the intensity fluctuations of the beam over a short period.
• Electrophoretic light scattering (ELS) measures the Zeta potential of a sample by passing a beam through a sample that has been induced to move using an electric field. The velocity of the particles and subsequent changes in frequency of the beam indicate the Zeta potential.
• Neither DLS nor ELS require calibration; instead, we recommend regular performance verification using sizing standards.
• Key considerations for DLS method development include measurement and attenuator positions, as well as the detection angle and sample concentration; for ELS, consider the concentration and the attenuator position.

Key quote

What we often get is ‘How do I calibrate my Zetasizer?’ And the answer is, ‘Well, you don’t need to, because it works off the first principles of physics.’

Reliable measurements without calibration

One of the advantages of using the Zetasizer Advance range for DLS and ELS measurements is that it does not require calibration. This is because both techniques work from the first principles of physics. Instead, our experts recommend regular performance verification using size standards purchased from Thermo Scientific. For DLS, these standards specify their hydrodynamic diameter, which you can then measure using 10 mmol of sodium chloride.

For ELS performance verification, Zeta potential transfer standards can be bought in batches, which you can use to verify that your measurements produce the same results that are written on the side of the syringe. The key for ELS performance verification is to ensure the cuvette is well-prepared in advance of the test by flushing it out with ethanol and then water to reduce the surface tension of the cell walls.

Top factors to consider in DLS and ELS method development

The right combination of settings is key to accurate DLS and ELS measurements. There are many parameters to consider here, including which cell you choose, and the dispersant used in the sample, but here we will highlight only a few. 

For DLS, the key things to consider are the measurement positions (where in the cuvette you will capture photons from) and the attenuator position, in other words, the amount of light that passes through the sample to allow for measurement. The vast majority of DLS measurements will be taken at a back angle, which enables you to make use of the non-invasive back scattering (NIBS) technique developed by Malvern Panalytical. This enables the measurement of a broad range of concentrations through the adjustment of the focusing lens position, thereby fine-tuning the experiment to fit the sample’s concentration.

Your attenuator position is also important when considering sample concentration. For instance, when measuring a low-concentration sample, you should use an attenuator that allows as much light in as possible to ensure you have enough signal to take a measurement. The Zetasizer Advance offers 11 attenuator positions for maximum flexibility.

For ELS, many of the same principles apply, however you should also pay close attention to the dilution of your sample. This is because Zeta potential is not only determined by the particles themselves, but also the medium in which they are suspended.

Key quote

"The Zetasizer Advance software has an array of automatic settings that are available to all users… However, in environments such as the pharma world, where you need to be compliant with certain regulatory bodies, it is often the case that many of these factors must be defined and locked down in the SOP."

Automated software for increased ease-of-use

There are many settings to choose from when setting up your experiment, and the Zetasizer Advance’s automation helps you set the best parameters to get the most accurate results possible. For example, the Zetasizer Advance can select the correct attenuator position for your DLS measurement based on the concentration of your sample. It can also choose an analysis model for your ELS measurement based on the conductivity of the sample.

Some regulatory bodies might require that you lock down such parameters in your standard operating procedures (SOP), however, these automatic settings are a helpful place to start when developing a measurement protocol. They can help you refine your methods and identify the settings that are most useful for your samples, allowing you to produce reliable, repeatable results. 

For more in-depth insight into fine-tuning your DLS and ELS techniques and how the Zetasizer Advance range makes it simpler, watch the webinar on-demand now!

Zetasizer Masterclass Webinar Series

Join us to learn about:

• Method development for DLS and zeta potential analysis
• The practical implications of USP 430 and 432
• Troubleshooting issues – with real examples from users

Take a look at the full Zetasizer Masterclass series of webinars here.