The power of orthogonal approaches to accelerate development of targeted therapies

01 Jan, 2023 | Guides & Resources
The power of orthogonal approaches to accelerate development of targeted therapies

A common challenge found in the world of scientific research is achieving recognition for topics not typically trending among influencers for the importance of fundamental research. Dr Georgia Atkin-Smith, a researcher at the Walter and Eliza Hall Institute and founder of the “Some Blonde Scientist” website noted in a recent post “most people think we should just fund the research that makes the drugs. But we obviously need to fund the research that makes the knowledge come to fruition before we can make the drugs.” Lipid Nanoparticles (LNPs) encapsulating mRNA have quickly gained popularity since used successfully during the COVID-19 pandemicby Pfizer and Moderna. However, the speed of vaccine development was only made possible because scientists had previously been working with this technology since the 1990s as drug delivery vectors for nucleic acid-based drugs and vaccines. The pandemic has demonstrated the need for developing fundamental knowledge that can be drawn upon in times of need. By using the right analytical tools and strategies we can ensure tomorrow’s therapeutic drugs and vaccines will be possible with today’s scientific breakthroughs. 

Shining a light on the drug development journey

The first step in the drug discovery and development journey involves identifying the molecules or compounds that bind to target proteins with the desired characteristics. This process however, comes with a high risk of failure -compounds may not behave as expected, lack the required activity, or show issues during development. Access to the best possible data can help researchers to make informed decisions about how the characteristics of the molecules and materials they are working with impact on their behaviour. In this way, a more complete picture of the interaction between lead molecules and target protein in terms of structure activity relationship and underlying interactions can be revealed. 

Advantage of using multiple analysis methods 

While proven analytical technologies can monitor a range of attributes, no one single biophysical characterisation technique can provide a complete picture, in the same way that no one single drug can be used to cure all types of diseases. Physicochemical characteristics like molecular size, composition, aggregation, surface charge, and structure can all have significant effects on the therapeutic molecule’s success. In addition, optimising one property may come at the detriment of another. These interdependent characteristics can impact stability during production, transport, storage, or affect the delivery of the drug in the body. From early drug discovery phases in the lab through to clinical, manufacturing and ultimately to the patient, adopting the power of orthogonal approaches of cutting-edge analytical solutions can ensure a more comprehensive understanding of the critical parameters needed that define the behaviour of a drug.

Analytical techniques are being increasingly implemented by biotech organisations

Access to accurate, robust, repeatable data is paramount to enable the rapid identification of optimal compounds that meet safety, bioavailability and likely processability requirements. 

For example, when formulating lipid nanoparticles (LNPs) as drug delivery vehicles, both the ratios of the ingredients and mixing technique must be optimised to ensure product consistency. The components of LNPs (cationic lipids, helper lipids such as distearoylphosphatidylcholine (DSPC), sterols such as cholesterol, and polyethylene glycol-containing (i.e., PEGylated) lipids) self-assemble during mixing so any small batch-to-batch variations like changes in flow rates can result in different compositions or LNP size. If the particles are too small they may be cleared from the body before they have a chance to take effect. On the other hand, larger particles can form aggregates and may fail to penetrate cells or induce unwanted side effects. Surface charge is also important as it can influence the uptake of LNPs to different cell types. 

Table 1 below lists a toolset of proven physicochemical analysis solutions helping researchers to make informed decisions about how the characteristics of the molecules and materials they are working with impact on their behaviour.

Table 1: Advanced physicochemical analysis solutions


Particle size


Particle concentration

Surface charge

Thermal stability

Higher order structure

Binding interaction

Particle composition












Multi detection SEC 













Particle size can influence dissolution, solubility, bioavailability and stability. The Mastersizer 3000 uses laser diffraction, one of the most widely used techniques for particle size distribution analysis. Suitable for both wet and dry samples, this highly repeatable (+/-1%) particle sizing technique delivers volume-based distributions in the size range 0.01 to 3500 microns and allows a specification developed in the laboratory to be transferred during scale-up. 

Two other well-established techniques used to monitor particle size and polydispersity include Dynamic Light Scattering (DLS) using the Malvern Zetasizerrange and NanoParticle tracking analysis (NTA) using the Malvern NanoSight range. Typically, DLS is used for rapid screening and tracking changes in size distribution. NTA provides high resolution size distribution and concentration measurements for individual nanoparticles while a fluorescence mode allows differentiation of fluorescing particles. Both methods measure changes in the scattering pattern of particles in suspension which is translated into size with larger particles diffusing more slowly than smaller particles.

Malvern Zetasizer Ultra is the most advanced system for the measurement of particle and molecular size, particle charge and particle concentration. Alongside the patented Non-Invasive Back-Scatter (NIBS) technology ideal for concentrated samples, the Zetasizer Ultra offers Multi-Angle Dynamic Light Scattering (MADLS®). Using back, side, and forward detection angles, MADLS offers a higher resolution than DLS, allowing it to identify additional populations of particles. It also enables calibration-free particle concentration analysis, resolving the individual concentrations of different size populations to provide even greater insight into samples. NTA offers even higher resolution but often requires sample dilution which can impact stability. The choice between these complementary techniques depends on factors, such as size and polydispersity, sample heterogeneity, and the product specifications.

Solubility issues during formulation can form undesirable aggregates which reduce the amount of active ingredient in the sample, thereby reducing efficacy, and they can stimulate immunogenic responses. Size Exclusion Chromatography (SEC) is a method commonly used to separate and measure the amount of different protein aggregate components and identify and characterise each of them by their molecular weight. But many proteins do not have globular structures making their measured molecular weights inaccurate using conventional SEC. 

The OMNISEC Multi-detection SECis a technique that combines the resolving power of chromatography with the revealing power of light scattering detectors and a viscometer. In a multi-detection SEC system, the light scattering detectors measure absolute molecular weight (as opposed to a column calibration system that can only provide a relative measurement) and the differential viscometer provides information about the molecular structure. This wider range of accurate data provides a more complete characterisation of the protein mixtures.


Differential Scanning Calorimetry (DSC) is a well-established label-free technique for structural characterisation and stability profiling of biomolecules and viruses in solution. DSC can analyse the effects of different storage conditions on the higher-order structure. It works by measuring the enthalpy (ΔH), temperature (Tm) and thermal stability of a vaccine. The MicroCal PEAQ-DSC system is the Gold standard stability assay platform which can be automated to support the generation of high integrity thermal stability data and deliver compliance with regulatory requirements.

Electrophoretic light scattering (ELS) is used in the characterisation and formulation development of products such as vaccines that use LNPs, liposomes and other nanoparticles as carriers, to determine size and colloidal stability. This method is useful in understanding what happens when a drug enters different cellular environments. The Malvern Zetasizer measures the zeta potential or apparent surface charge of particles and molecules, indicating sample stability and/or propensity to aggregate. The apparent surface charge can be used to detect formulation condition differences.


Understanding the binding affinity, or the interaction between biomolecules, is key for the design of drugs that bind their targets selectively and specifically. Grating-Coupled Interferometry (GCI)and Isothermal Titration Calorimetry (ITC) can be used together to provide highly quantitative affinity (KD) values. Both are label-free quantification techniques, allowing the use of native molecules. GCI is an optical method that measures the change in refractive index on a sensor surface caused by the binding event and is used to study the affinity and kinetics of an interaction. The Creoptix WAVE biosensor with proprietary GCI technology measures KD values in the millimolar to picomolar range and additionally determines the kinetics of an interaction, more specifically, the on (ka) and off (kd) rates. It an aid screening for real-time label-free binding kineticsusing low sample volumes, even from unpurified material. The MicroCal ITC range measures the heat change associated with the binding event. It measures KD values in the millimolar to nanomolar range and determines the binding stoichiometry and binding thermodynamics of the interaction. Both kinetics and thermodynamics are important in the characterisation of intermolecular interactions.

At ATA Scientific, we’re proud to enable scientists to accelerate Drug Discovery by providing cutting-edge tools for molecular interaction analysis. If you need assistance in determining if your application is possible, contact us.

For further details contact 
ATA Scientific Pty Ltd
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