Protein Characterization

Offers a range of analyses and studies to support the characterization of proteins, including differential scanning fluorimetry, dynamic light scattering, and viscosity measurement.

We offer a comprehensive range of bioanalytical methods to support your drug development needs


Our services include determining protein identity, purity, content, and bioactivity using high-throughput technologies. We work with a wide range of proteins in different formats, including purified proteins, protein-ligand complexes, and protein formulations. HTD is equipped to provide highly detailed and accurate characterization of proteins for research and development purposes for our clients.

Protein Aggregation 
We use various orthogonal techniques for characterizing protein aggregation. These techniques include size exclusion chromatography, dynamic light scattering and flow cam. These methods allow us to detect, quantify and understand the formation of protein aggregates in solution, which can affect the activity, stability, and safety of proteins. By understanding the conditions and factors that lead to aggregation, we can help our clients develop strategies to minimize or eliminate aggregation in their protein-based products.


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Determination of Particle Size

Dynamic light scattering (DLS) is a technique that is used at HTD Biosystems to study particle size and aggregation behavior of macromolecules and nanoparticles. We can measure the particle size either under isothermal condition for example at 20C or as the temperature is changing under a controlled ramp. For proteins, we also offer KD studies to evaluate protein-protein interaction and solution stability.

Determination of Particulate Matter 

Subvisible particle characterization by Flowcam microscopy is a technique that is used to evaluate the size, shape, and distribution of subvisible particles in a sample. Subvisible particles are particles that are smaller than visible particles, typically ranging in size fromin  1 micrometer to 50 micrometers size range, and they can be found in a variety of products, including injectables. 

USP has strict guidelines for acceptable limits of particulate matter in parenteral drug products as described in US Pharmacopiea (USP <787> Subvisible Particulate Matter in Therapeutic Protein Injections, USP <788> Subvisible Particle Matter in Injections, and USP <789> Particulate Matter in Ophthalmic Products.  
We have discussed the use of FlowCam in protein characterization and formulation in our recent interview with Fluid Imaging .

Determination of Protein Identity & Purity

To establish protein identity and purity, we use protein chip bioanalyzer technology, which utilizes high-throughput SDS-PAGE to determine the molecular weight of protein monomers and detects any impurities or aggregates.  Additional methods include HPLC and UPLC.

Determination of Protein Content

For accurate determination of protein content, our scientists employ a combination of techniques including UV spectroscopy and HPLC. UV spectroscopy measures the absorption of ultraviolet light by a protein to calculate its concentration in a solution. To further confirm the results, we also utilize High Performance Liquid Chromatography (HPLC), in partnership with other organizations. By combining these methods, we can ensure a precise and accurate determination of protein content.

Differential scanning fluorimetry (DSF)

Differential scanning fluorimetry (DSF) is a technique that is used to study protein conformation, unfolding and thermal denaturation of proteins. DSF measures the changes in the inherent fluorescence emitted by a protein as it is heated without addition of any dyes. 
Using DSF, we can determine the temperature at which a protein starts to denature (the onset of melting temperature, or Tm-onset), as well as Tm (denaturation temperature) to determine the stability of the protein in different formulations. 
Recently we collaborated with NanoTemper on the use of nDSF in protein formulation development using our iFormulate platform.  

Viscosity Measurement

Viscosity measurement is a useful tool for understanding the rheological properties of proteins and other biomolecules, and for developing stable protein formulations for drug development. Viscosity is critical quality attribute (CQA) for high concentration protein formulations that need to be below 20 cPa for subcutaneous injections.


Osmolality Measurement

Osmolality is also a CQA for subcutaneous administration and bolus intravenous injections of drug products.  We measure osmolality using the freezing point method at HTD Biosystems.  This technique provides a rapid and accurate measurement of solution osmolality in 50 ul sample, reported in mOsmol/kg.  It is utilized during our formulation screening, optimization, and quality control testing.