Particle stability determines whether a colloidal system stays uniform over time or begins to degrade through processes like aggregation or sedimentation. In industries such as pharmaceuticals, food, and coatings, dynamic light scattering (DLS)—commonly performed using instruments like the Malvern Panalytical Zetasizer —is widely used to monitor particle size and detect early signs of instability.
Maintaining stable particles is important for ensuring consistent product performance, reliable drug delivery, and preserving properties such as texture, efficacy, and shelf life. DLS provides a fast and simple way to track changes in particle size, helping identify when a formulation may be starting to break down.
This article explains the basics of dynamic light scattering, the key parameters it measures, and how these can be used to assess and improve particle stability.
How Dynamic Light Scattering Works
Dynamic Light Scattering (DLS), sometimes referred to as Photon Correlation Spectroscopy (PCS) or Quasi-Elastic Light Scattering (QELS), is a simple, non-invasive technique used to measure the size of particles in liquids, typically at the nanoscale. In a DLS measurement, the sample is placed in a cuvette and illuminated with a laser. As the particles move randomly due to Brownian motion, the scattered light fluctuates in intensity. By analysing these fluctuations using autocorrelation, the system determines how fast the particles are moving (their diffusion rate). This information is then used to calculate particle size using the Stokes–Einstein relationship [1].
Brownian Motion and Light Scattering
Particles suspended in a liquid are constantly moving due to collisions with surrounding solvent molecules—this random movement is called Brownian motion. When a laser shines on these particles, the light they scatter changes in intensity over time. Smaller particles move faster and cause quicker fluctuations, while larger particles move more slowly. By analysing these fluctuations, DLS determines how fast the particles are moving and uses this information to calculate their size through the Stokes–Einstein relationship.
In DLS, particle size is reported as the hydrodynamic diameter, which describes how a particle moves in a liquid rather than just its physical core size. This means the measurement includes not only the particle itself, but also any surface coatings and the surrounding layer of solvent molecules. As a result, DLS sizes are often larger than those measured by imaging techniques like Transmission Electron Microscopy, which only show the dry particle core. In simple terms, DLS tells you how a particle behaves in solution, while imaging shows what it looks like in a static state [2].
From Autocorrelation to Particle Size
A correlator is basically a signal comparator designed to measure the degree of similarity between two signals, or one signal with itself at varying time intervals. The correlator used in a DLS instrument will construct the correlation function G(τ) -a mathematical description of how similar the scattered intensity is to itself at different time delays – of the scattered intensity:
G(τ) = <I(t).I(t+τ)> Where τ =the time delay.
The shape of the correlation function reveals key information about the properties of your sample.
Small particles diffuse rapidly, producing a fast decay, while larger particles move more slowly and yield a slower decay. A monodisperse sample shows a single-exponential decay, whereas a polydisperse sample produces a multi-exponential decay due to the presence of different particle sizes. From the decay rate, the translational diffusion coefficient (D) is obtained. This is converted to the hydrodynamic diameter using the Stokes–Einstein equation:
where k is the Boltzmann constant, T is absolute temperature, and η is the viscosity of the solvent. [3]
The intercept of the correlation function (at ) is a key data quality indicator. Values close to 1 indicate good data quality. Lower intercepts suggest interference such as dust, fluorescence, or multiple scattering (often called flare). These effects can often be reduced by lowering sample concentration or adjusting the measurement position—for example, measuring closer to the cuvette wall in backscatter mode (Non-Invasive Back Scatter). Conversely, unusually high intercepts (greater than 1) may indicate significant number fluctuations, where the number of particles within the laser beam is changing significantly over time [4].
Key DLS Outputs and What They Mean
Dynamic light scattering (DLS) measurements generate several parameters that describe particle size and sample quality. Correct interpretation requires understanding what each parameter truly represents—and its limitations. Misreading DLS data can easily lead to incorrect conclusions about particle size, distribution, or sample stability, particularly for complex or polydisperse systems.
Z-Average and Cumulant Analysis
DLS reports particle size distributions, typically as intensity-, volume-, and number-weighted plots. The primary output is the Z-average (cumulants mean), an intensity-weighted mean hydrodynamic diameter derived from the autocorrelation function. The intensity-weighted distribution is the direct measurement and strongly emphasises larger particles, as scattering intensity scales approximately with the sixth power of particle diameter () for Rayleigh scatterers. This means that even a small number of large particles or aggregates can dominate the intensity distribution. As such, the Z-average is most reliable for monodisperse or narrowly distributed samples and is often used for quality control and comparison purposes [5].
Volume weighted (d3 or mass) and number weighted (d0) distributions are calculated from the intensity data to better reflect the relative proportions of material or particle counts. However, these transformations rely on assumptions about the form of the particle and its physical properties (RI) and can introduce uncertainty, particularly for polydisperse samples. A common misinterpretation is to assume that a narrow number-weighted distribution indicates the absence of large particles. In reality, such particles may still be present but underrepresented. For this reason, intensity data should always be considered alongside derived distributions [6].
Polydispersity Index (PDI)
The polydispersity index (PDI) provides a measure of the width of the particle size distribution. PDI ranges from 0 (perfectly monodisperse) to 1 (highly polydisperse). The PDI provides a useful, though simplified, measure of the breadth of a particle size distribution. As a general guide, values below 0.1 indicate a narrow, monodisperse system; 0.1–0.25 suggests a moderately narrow distribution; and values above 0.5 are typically associated with broad, multimodal, or aggregated samples. In pharmaceutical applications—particularly for nanoparticle formulations such as lipid nanoparticles (LNPs)—regulatory expectations often target a PDI below 0.2. PDI is therefore considered a critical quality attribute, as increases during storage can serve as an early warning sign of aggregation, often before any significant change in average particle size is observed. However, it is important to recognise the limitations of PDI: because it is derived from cumulant analysis, it assumes a single, unimodal distribution and does not reliably describe systems with multiple particle populations. In such cases, more detailed size distribution analysis or orthogonal techniques such as nanoparticle tracking analysis (NTA) are required for accurate characterisation [7].
Intensity, Volume, and Number Distributions
Beyond Z-average and PDI, DLS instruments can display particle size distributions as intensity-, volume-, or number-weighted plots. The intensity distribution is the primary measurement and should always be examined first, as it is most sensitive to larger particles; even a small secondary peak at high sizes can indicate aggregation, despite a normal Z-average. Volume and number distributions are derived from the intensity data and can help visualise relative proportions, but they rely on assumptions and are prone to error—particularly at the distribution tails. As a result, transformed distributions should be interpreted cautiously and always alongside the intensity distribution for an accurate assessment of sample quality [7].
Zeta Potential as a Colloidal Stability Indicator
Zeta potential is measured by electrophoretic light scattering (ELS), which is often integrated into the same instrument (e.g. Zetasizer). While DLS measures particle size based on Brownian motion, ELS measures particle mobility in an applied electric field to determine zeta potential, a key indicator of surface charge and colloidal stability. Together, DLS and ELS provide a more complete understanding of a system, linking particle size with the electrostatic interactions that influence aggregation and stability.
How Zeta Potential is Measured
In ELS, an oscillating electric field is applied to the sample causing charged particles to migrate toward the electrode of opposite charge. This movement is tracked using Laser Doppler velocimetry, which measures the particle speed as electrophoretic mobility. The mobility is then converted to zeta potential using Henry’s equation, typically applying the Smoluchowski approximation for aqueous systems or Hückel approximation for non-polar systems. Advanced tools such as the patented M3-PALS (Mixed Mode Measurement Phase Analysis Light Scattering) method, used in the Malvern Panalytical Zetasizer series, enable accurate zeta potential measurements across a wide concentration range. Because electrophoretic mobility depends on solvent viscosity, temperature must be carefully controlled and reported to ensure reliable and reproducible results [8].
Interpreting Zeta Potential for Stability
Colloidal stability is often interpreted through DLVO theory (Derjaguin, Landau, Verwey, and Overbeek), which describes stability as the balance between electrostatic repulsion and van der Waals attraction. As a general guideline, absolute zeta potential values greater than ~30 mV indicate good electrostatic stability, while values below ~10 mV suggest a higher risk of aggregation. However, these thresholds apply primarily to electrostatically stabilised systems, such as charged polymer or lipid nanoparticles. Sterically stabilised systems—such as PEGylated nanoparticles—can remain stable even when the zeta potential is close to zero, so applying electrostatic rules in these cases can be misleading. It is also important to recognise that zeta potential is highly sensitive to experimental conditions, including pH, ionic strength, and formulation components, and these parameters should always be reported alongside the measurement [9].
Designing a DLS Stability Study
Rigorous stability studies require systematic design — choosing appropriate stress conditions, time points, measurement parameters, and statistical approach. DLS and zeta potential measurements are essential in nanoparticle stability protocols providing complementary insight into changes in particle size, distribution, and colloidal stability over time. When properly designed, these studies can detect early signs of instability—such as aggregation or surface changes—before they become evident through visible or functional changes [9].
Stress Conditions and Time Points
A well-designed study typically combines real-time and accelerated conditions. Real-time stability involves storing samples under intended conditions and measuring at defined intervals to monitor changes under realistic use scenarios. Accelerated stability studies expose samples to elevated temperatures (such as 25 °C, 37 °C, or 40 °C) to predict long-term behaviour in a shorter timeframe. Additional stress testing is often essential: freeze–thaw cycling is particularly critical for frozen formulations, including mRNA lipid nanoparticles, where repeated freezing and thawing can induce aggregation or structural disruption. Agitation or mechanical stress studies may also be included to simulate handling, transport, or administration conditions.
At each time point, key parameters should be consistently measured, including Z-average (mean particle size), polydispersity index (PDI), and zeta potential. Where relevant, functional attributes such as encapsulation efficiency should also be assessed using orthogonal techniques such as Nanoparticle tracking analysis (NTA) technology with fluorescence detection used in the Malvern Panalytical NanoSight Pro system. Together, these measurements provide a comprehensive view of both physical and functional stability [9].
Measurement Best Practice
Careful sample preparation and consistent measurement practices are essential for reliable DLS stability studies. Samples should be appropriately diluted in a buffer that matches the formulation’s ionic strength to avoid altering particle behaviour, and, where necessary, filtered (e.g. using a 0.2 µm syringe filter) to remove dust. Prior to measurement, samples should be allowed to equilibrate to the set temperature within the cuvette to ensure stable and reproducible conditions. Good practice also includes performing multiple measurement replicates and preparing independent samples to assess variability and improve data confidence. Reporting should follow established standards such as ISO 22412:2025, including key parameters like Z-average, PDI, temperature, solvent viscosity, refractive index, and measurement settings [10].
Temperature-controlled measurements using the Zetasizer Advance range — comprising the Zetasizer Lab, Zetasizer Pro, and Zetasizer Ultra — allow stability studies to be conducted across a defined temperature ramp, with automated data collection enabling rigorous time-point measurements without manual intervention. The Zetasizer Ultra adds Multi-Angle Dynamic Light Scattering (MADLS) capability, providing angle-independent size measurements that are particularly valuable for polydisperse samples or those with complex scattering signatures. Measurements can be made from just 1 µL of material with the Zetasizer low-volume sizing cell, ideal for scarce or high-value samples [11, 12].
- Zetasizer Ultra product page
- Zetasizer Pro product page
- Zetasizer Lab product page
- Multi-Angle Dynamic Light Scattering (MADLS)
Applications Across Research Fields
DLS and zeta potential are applied across a broad range of colloidal systems. The technique’s speed, sensitivity, and non-destructive nature make it well-suited to both early-stage formulation screening and ongoing quality control.
Pharmaceutical Nanoparticles and Lipid Nanoparticles
The Zetasizer plays a pivotal role in the development and manufacturing of pharmaceutical and lipid nanoparticles because it enables precise measurement of key physicochemical properties that define product quality and stability. By using advanced light scattering and electrophoretic techniques, the instrument provides accurate data on particle size, polydispersity, and zeta potential—all critical quality attributes that influence formulation performance and therapeutic efficacy. These measurements help researchers evaluate nanoparticle uniformity, monitor aggregation, and assess electrostatic or steric stability, ensuring that formulations remain consistent from lab-scale development to large-scale production. As highlighted in the recent Malvern Panalytical ebook “Overcoming Challenges in LNP Vector Analysis: Key Tools, Techniques, and Considerations”, comprehensive characterisation of lipid nanoparticles requires accurate assessment of size, dispersity, and surface charge, as these directly impact stability, delivery efficiency, and regulatory compliance. When paired with complementary technologies such as the NanoSight Pro, the Zetasizer allows for a comprehensive characterisation approach that informs better process control and faster optimisation of mRNA-LNPs and other novel drug delivery systems [13].
Protein Therapeutics and Biologics
Dynamic light scattering (DLS) using the Zetasizer Advance plays a critical role in protein therapeutics and biologics by enabling rapid, reliable characterisation of protein size distributions and aggregation states—parameters that directly impact efficacy, safety, and reproducibility. As demonstrated in an application note on amyloid fibrils, DLS can quickly measure key metrics such as Z-average size and polydispersity index (PDI) within minutes, allowing researchers to optimise processing conditions to produce uniform protein structures of a defined size (~50 nm) suitable for biological function. This capability is particularly important in biologics, where subtle changes in aggregation or size heterogeneity can alter bioavailability, immunogenicity, and therapeutic performance. Zetasizer using DLS provides a fast, complementary alternative to high-resolution techniques like TEM or AFM, enabling routine monitoring of protein formulations during development and manufacturing. By ensuring consistent particle size and detecting aggregation early, Zetasizer-based DLS supports robust process optimisation, improves reproducibility in cell-based assays (such as neuronal uptake and seeding), and ultimately helps ensure the quality and stability of protein therapeutics [14].
Environmental and Food Science
Dynamic light scattering (DLS) with instruments such as the Zetasizer Advance is increasingly important beyond pharmaceuticals, playing a critical role in environmental, food, and agricultural systems where colloidal behaviour governs performance and safety. In water treatment and environmental monitoring, DLS enables the detection and characterisation of contaminants—including microplastics and engineered nanoparticles—by measuring their size distribution and aggregation behaviour in complex aqueous matrices, helping assess transport, fate, and potential ecological impact. This capability is equally valuable in food science, where the stability of emulsions (e.g., dairy products, beverages, and sauces) depends strongly on droplet size distribution and resistance to coalescence; DLS provides rapid, non-invasive measurements that directly relate to texture, shelf life, and product consistency. In agriculture, nano-formulations such as nano-pesticides and fertiliser carriers rely on controlled particle size and dispersion stability to optimise delivery efficiency and minimise environmental loss, making DLS essential for formulation development and quality control. Overall, by enabling fast, accurate assessment of particle size, dispersity, and aggregation in complex colloidal systems, Zetasizer-based DLS supports improved process understanding, environmental safety, and product performance [15].
Australian Research Spotlight
UNSW:
At the University of New South Wales, the RNA Institute and Flow Cytometry Facility hosts both Zetasizer Nano ZS and Zetasizer Ultra systems, which are actively used for nanoparticle and biomolecular characterisation. The primary users are facility scientists who oversee instrument access, training, and method development. Their work frequently involves extracellular vesicles, protein systems, and biological colloids, meaning the Zetasizer is embedded in workflows spanning biomedical research and nanomedicine. Because the facility operates as a shared resource, these staff members also indirectly support a broad base of PhD students and postdoctoral researchers who rely on DLS and zeta potential measurements.
Griffith University:
At Griffith University, the Queensland Micro and Nanotechnology Centre provides access to a Zetasizer Nano ZS within its research infrastructure. Instrument use is closely associated with researchers linked to nanotechnology and colloid science applications. The system is routinely applied in studies of nano-emulsions, drug delivery systems, and protein formulations, indicating consistent demand from both materials science and biopharmaceutical research groups. As with most shared facilities, the Zetasizer, supports a number of users, particularly postgraduate researchers conducting formulation and stability studies.
University of Sydney:
Within University of Sydney, the Sydney Analytical platform has installed their Malvern Panalytical Zetasizer as an important part of its scattering and diffraction capabilities. Facility specialists support researchers using DLS for particle sizing and zeta potential analysis. Zetasizer is usually the first instrument most users start with for studying particles in solution. The instrument is widely applied across disciplines including chemistry, materials science, and life sciences, particularly for nanoparticle characterisation and colloidal stability assessments. Its integration into a central analytical hub means it is accessed by a diverse research community.
Curtin University:
At Curtin University, School of Molecular and Life Sciences chemistry facilities, the Zetasizer is used by researchers in mineral processing, bionanomaterial synthesis, agriculture and food science and various colloidal systems to study particle aggregation and surface charge behaviour. The instrument is embedded within laboratory-level workflows, with frequent use by PhD candidates and research staff providing essential size and stability measurements to understanding life in all its complexity – from molecules to ecosystems and global systems.
University of Technology Sydney (UTS):
UTS has installed their Zetasizer instruments at two facilities: Microstructural Analysis Unit (MAU) and the Centre for Technology in Water & Wastewater (CTWW) facility. The MAU is a centralised large equipment facility which provides researchers in both the physical and biological sciences with access analytical microscopy tools as well as materials analysis and fabrication equipment. The CTWW environmental engineering labs facilitate high impact water and wastewater research. These laboratories house world-class equipment including the Zetasizer DLS system, which is shared with a number of other universities and research organisations.
University of Queensland:
The Centre for Microscopy and Microanalysis provide a suite of analytical instrumentation including the Zetasizer Ultra and a high standard of training programs for university researchers. The Australian Institute for Bioengineering and Nanotechnology (AIBN) is one of Australia’s leading nanoparticle and biomaterials research institutes active in LNP development for gene therapy and vaccine delivery. The BASE facility supports mRNA-LNP formulation research. DLS-based size and stability characterisation is central to formulation workflows.
Monash University:
Researchers at the Hybrid Assembly use light scattering and microscopy techniques to investigate the micro- and nano-structure of the materials, where the Zetasizer is key. At the school of Chemical and Biological Engineering studies are focused on pollution control, energy management, and transformation of waste into useful products. This incorporates such projects as; bioplastics from paper and food waste, recycling tyres to generate fuel and implementing wastewater processes to create clean drinking water. Monash University Institute of Pharmaceutical Sciences (MIPS) conducts formulation research including nanoparticle drug delivery. Melbourne Centre for Nanofabrication uses their Zetasizer study of organic and inorganic nanoparticles, ceramics, pigments and inks,
emulsions and pharmaceuticals.
Adelaide University:
At the University of Adelaide, researchers within the Future Industries Institute and other science labs utilise Zetasizer systems as a core tool for characterising nanoscale materials and colloidal systems. The Zetasizer is used for dynamic light scattering (DLS) and zeta potential measurements in projects spanning nanomedicine, advanced materials, and environmental technologies. Typical use cases include assessing nanoparticle size distributions in drug delivery systems like virus‐like particles (VLPs), evaluating dispersion stability in polymer and coating formulations, and monitoring surface charge behaviour in water treatment and membrane research. It supports a wide range of users—from PhD researchers to senior scientists—providing rapid, reproducible insights into particle behaviour that are critical for both fundamental studies and applied industrial translation [16].
These contributions from are just some examples from the Australian research community currently harnessing the capabilities of Zetasizer – based DLS. It reflects a strong capability in translational nanomedicine, supported by access to high-performance characterisation instrumentation at research infrastructure facilities across the country.
Advantages and Limitations of DLS and Complementary Techniques
DLS limitations
Malvern Zetasizer Dynamic light scattering (DLS) is widely used because it provides rapid, non-destructive measurement of particle size and polydispersity in liquid dispersions. A key advantages is speed—measurements typically take minutes with minimal sample preparation. DLS is highly sensitive to nanoscale particles (from <1 nm up to a few microns) and can deliver intensity-weighted size distributions, making it ideal for detecting aggregates or small changes in stability. The Zetasizer can measure zeta potential, offering insight into surface charge and colloidal stability in a single platform. However, like all methods, DLS has important limitations: results are strongly biased toward larger particles because scattering intensity scales with the sixth power of particle diameter. This means even a small number of aggregates can dominate the signal and mask the primary population. It also assumes spherical particles and struggles with highly polydisperse or multimodal samples, where deconvolution of size populations becomes challenging.
The Zetasizer Ultra overcomes common DLS challenges with its patented multi-angle dynamic light scattering (MADLS) and Non Invasive Back Scattering (NIBS) technology, which enables the measurement of particle size and concentration with improved resolution and accuracy. Adaptive Correlation reduces noise and delivers accurate data up to three times faster than traditional DLS. Zetasizer low volume sizing cell is ideal for measuring data from as little as 1 µL, making it suitable for scarce or high-value materials. Combined with Zetasizer Sample Assistant and OmniTrust for data integrity, its capabilities are enhanced further, ensuring reproducible results even with complex samples or when users are not light scattering experts [17].
When to use complementary techniques:
- Nanoparticle Tracking Analysis (NanoSight Pro): Better for polydisperse samples, provides particle-by-particle size and concentration measurement
- Transmission Electron Microscopy (TEM/cryo-TEM): Direct visualisation of particle morphology and core size
- Asymmetric Flow Field-Flow Fractionation (AF4): High-resolution separation before size measurement
For samples where DLS alone is insufficient — such as highly polydisperse formulations or samples requiring particle concentration data — Nanoparticle Tracking Analysis using the NanoSight Pro provides particle-by-particle size distribution and direct count data, complementing DLS measurements from the Zetasizer to provide a more complete characterisation picture [17].
ATA Scientific’s Commitment to Particle Characterisation Research
ATA Scientific supports scientists within Australia and New Zealand with a range of complimentary particle characterisation instrumentation, including the full Zetasizer Advance range and NanoSight Pro for NTA measurements. With over 35 years serving the Australian research and industrial community, our technical team provides support from instrument selection through to measurement optimisation and data interpretation. Contact us for a demonstration or for hands-on training using the Malvern Panalytical Zetasizer.