Biochemical Assays
Biochemical assays are essential to generate the reliable, quantitative data required to select the right targets, validate mechanisms, and de-risk early drug discovery programs. When these assays are well designed, they reduce experimental noise, improve hit quality and help avoid costly re-work later in the pipeline.
Our integrated biochemical assay platform spans recombinant protein expression and purification, HTS-ready functional assay development, and tailored secondary assays.
This end-to-end approach accelerates hit identification, strengthens hit-to-lead decision-making, and increases the efficiency of lead optimization.
Protein
production
Functional HTS assay
development
Protein production and
assay adaptation for HTS/
compound screening
Hit-to-Lead
Types of Biochemical Assays we develop
Robust, scalable, and high-quality biochemical assays are developed at Axxam to seamlessly support HTS and downstream drug discovery.
Within this structured development workflow, we offer a broad suite of assay technologies:
- Enzymatic assays to accurately measure enzyme activity across diverse target classes.
- Proximity assays for studying protein–protein, protein–nucleic acid, and protein–small molecule interactions, including PROTAC mechanisms.
- Biophysical binding assays that deliver precise interaction data and are fully compatible with HTS workflows.
Enzymatic assays
Our advanced enzymatic assay development addresses a wide range of target classes, including:
Oxidoreductases
– Histone demethylases
– Dioxygenases
– Metabolic enzymes (Warburg effect)
Transferases
– Protein kinases (Ser, Thr, Tyr)
– Nucleotide kinases
– Metabolic enzymes (Warburg effect)
Hydrolases
– Peptidases (Asp, Cys, Ser, Met, Thr)
– Lipases
– HDACs
– Phosphatases
– DNA/RNA/Nucleotide hydrolases
– Phosphodiesterases
– Deubiquitinases
Lyases
– Metabolic enzymes (Warburg effect)
Isomerases
Ligases
In-house libraries of surrogate substrates for protein kinases, proteases, and esterases enable detailed analysis of enzyme selectivity and support the identification of optimal target-specific substrates, improving hit quality in discovery programs.
Each enzymatic assay is thoroughly characterized and optimized through the determination of kinetic parameters, assessment of the stability of key assay components, and validation using tool reagents and reference compounds. In addition, their robustness for compound testing is carefully evaluated under workstation-assisted HTS conditions, ensuring reliable and reproducible screening result.
Proximity assays
Understanding how biomolecules interact is central to many early drug discovery programs, especially when targeting complex signaling or degradation mechanisms. For this reason, proximity and cell-free interaction assays provide a powerful way to investigate these interactions under fully controlled conditions, from simple binary systems to complex ternary and higher-order assemblies.
For biochemical proximity assays, we typically use TR-FRET or fluorescence polarization readouts. As a result, these assays achieve high sensitivity, reliability, and full compatibility with HTS automation.
A selection of interaction formats developed in recent case studies includes:
- Protein-protein interaction
- Protein-peptide interaction
- Protein-DNA interaction
- Protein-RNA interaction
- RNA-targeting compounds
- Protein-compound interaction e.g., PROTACs, molecular glues
These cell-free interaction assays have contributed to successful drug discovery programs focused on modulating protein and RNA dynamics. Applications include the discovery of Proteolysis-Targeting Chimeras (PROTACs), Ribonuclease-Targeting Chimeras (RIBOTACs), protein or RNA stabilizers, and molecular glues designed to restore or enhance physiological cellular turnover.
Before entering screening campaigns, each proximity assay is rigorously optimized and validated to meet HTS requirements. First, our biochemical team determine affinity parameters and assess the stability of key assay components. Next, they validate performance with tool reagents and reference compounds. Finally, automated HTS platforms confirm robustness for compound screening, ensuring reliable, reproducible, and high-quality screening data.
Biophysical binding assays (TSA/MST)
A critical step in any drug discovery program is obtaining clear and reliable evidence that a compound directly interacts with its target. To address this need, we provide a comprehensive platform to study compound–target interactions in solution using Thermal Shift Assay / Differential Scanning Fluorimetry (TSA/DSF).
This biophysical method detects ligand-induced changes in target thermal stability and does not require immobilization. As a result, it is well suited for early screening and target validation.
TSA/DSF assays are fully HTS-compatible and run in 384-well format on automated screening platforms. Dedicated data-analysis software ensures throughput comparable to standard high-throughput screening workflows.
To support challenging targets, we also offer an in-house developed dye library and a dedicated optimization platform enable reliable TSA/DSF assay setup even for proteins that perform poorly under standard conditions. In addition, this technology can be successfully applied not only to protein ligands but also to compounds interacting with nucleic acids such as RNA, broadening its applicability in modern drug discovery.
The TSA/DSF platform is complemented by Microscale Thermophoresis (MST) and Temperature-Related Intensity Change (TRIC). These biophysical assays enable precise determination of protein–ligand binding affinities in solution under mid-throughput conditions and are well suited for characterizing compound series and screening targeted libraries and fragment collections.
HTS-Compatible readouts and formats
Cell-free assays are configured in 384- or 1536-well formats, while assays transferred by clients can be miniaturized from any previous setup.
We apply a broad range of HTS-compatible readouts and state-of-the-art screening technologies, including:
- Fluorescence-based methods
– Fluorescence intensity
– Time-resolved fluorescence (TRF)
– FRET and TR-FRET
– Fluorescence polarization - Absorbance
- Luminescence
- AlphaScreen®
- Radiometric systems
– FlashPlate® technology
– Scintillation Proximity Assay (SPA)
- Fluorescence-based methods
In addition, we evaluate alternative assay types and recombinant target variants in parallel using a matrix approach during the proof-of-principle phase.
This strategy identifies the assay combination with the highest performance, which is then shortlisted for HTS.
As a result, each program benefits from enhanced assay robustness, reproducibility, and effectiveness in the downstream drug discovery workflow.
Recombinant target proteins and production platforms
Biochemical assays at Axxam can be developed starting from in-house purified recombinant proteins, client-provided targets, or commercial protein sources. Our team has extensive expertise in producing cytosolic, secreted, and transmembrane proteins using the most reliable expression platforms, including:
- Bacterial systems
- Insect cells with the baculovirus expression system
- Mammalian high-density suspension cultures
Whenever feasible, we express full-length enzymes to preserve all relevant regulatory domains. This approach maintains both orthosteric and allosteric binding sites and supports the identification of physiological modulators and potential inhibitors.
In addition, we express high-molecular-weight targets, including heteromultimeric protein complexes, to enable their direct use in HTS-compatible biochemical assays.
Why develop a biochemical assay with us
Developing a biochemical assay requires more than setting up a test. It demands precision, scalability, and direct relevance for drug discovery decisions. Axxam addresses these challenges by combining deep scientific expertise with advanced technologies to deliver assays that meet the highest standards of quality and reliability.
Proprietary recombinant protein technologies ensure consistent target production, while custom assay design is tailored to specific targets and screening requirements. In parallel, HTS-ready solutions are optimized for 384- and 1536-well formats to enable seamless scale-up.
Through a proven end-to-end workflow, Axxam transforms complex assay development into robust, reproducible, and cost-effective screening solutions. As a result, hit identification is accelerated, hit-to-lead decisions are strengthened, and lead optimization becomes more efficient.