Overview for Affinity & Kinetic Characterization
The establishment of the rate of interactions between two or more molecules and their rate of dissociation is key to understanding biological molecules complex formation and stability; two factors that are important in elucidating ligand-receptor binding mechanisms and stability during the development of biologics drug molecules. Two systems with the same affinity may exhibit completely different binding mechanisms, hence the evaluation of the affinity constants in conjunction with the association and dissociation rates of binding is crucial in the selection process for optimal drug candidates. While ELISA has been the dominant technique in affinity characterization of biomolecules, real-time label free technologies have become increasingly popular due to their ability to provide more detailed information in the kinetics of binding of biomolecules.
Large molecules kinetics characterization
The BLI Octet® family of instruments accurately measures kinetic constants by bringing the detection surface directly to the sample, eliminating the need for microfluidics. This unique approach utilizing label-free, real-time analysis streamlines laboratory workflow and expedites assay development. It allows direct measurement of crude samples while minimizing instrument maintenance.
Fc receptor binding assays on the Octet system
The safety and efficacy of a therapeutic monoclonal antibody can be greatly impacted by its ability to bind to both the target and to the FcγR. Antibodies are often selected based on their binding properties to FcγRs. They are sometimes engineered to achieve desired FcγRs binding properties. As a result, the assessment of binding affinities of these antibodies to FcγRs is an integral part of biotherapeutic development processes.
- Octet systems offer high throughput and sensitive methods for Fc receptor binding analysis
- A variety of biosensor surfaces are available and allow for flexibility and rapid optimization of assay format
Protein - Small molecules and peptide kinetics & affinity characterization
Small molecule and peptide therapeutic drugs are highly sought after in most areas of disease research due to their desirable pharmacological properties and low propensity for immunogenicity. In small molecule drug discovery, the path to lead molecules can stem from many sources or starting points including fragment screening, high throughput screening, de novo structural design, etc. The determination and evaluation of the affinity of small molecule binding to a therapeutic target is a significant component of the drug discovery process and lead optimization. The hit-to-lead and lead optimization process are essential to accurately determine biological potency in vitro so that structure-activity relationships (SAR) can be used for efficient structural design. Learn how Octet RED96e, Octet RED384, Octet HTX and Pioneer SPR platforms can be used to characterize small molecule and peptide systems.
Characterization of high affinity biologic interactions
Target binding characterization is an essential analytical step for the selection of high affinity (KD <1 nM) and highly specific biologics regardless of the types of molecules. A kinetic analysis further describes the components of association and dissociation that comprise the overall affinity interaction. For example, two lead compounds may possess a similar affinity (KD) to its target, but their differences in kinetic rate constants of association and dissociation can be used to estimate which will be more useful in vivo. Accurate analysis of these kinetic rate constants is therefore important information for lead selection and predicting the efficacy of protein therapeutics. The Pioneer SPR system with next generation SPR injections improves the efficiency of the characterization process over traditional SPR by determining the kinetics and affinity in a single step. The next-generation OneStep® gradient injection featured on the Pioneer platform dramatically increases the speed of affinity characterization while maintaining accuracy and high confidence in results.
Resources of Affinity & Kinetic Characterization
Application Note Commitment to Covalency : Kinetics of Irreversible Inhibitors on the Pioneer FE System
The principal role of assay groups in drug discovery is to provide reliable methods, analysis, and data for confident decision-making about series progression. Particular assays are chosen to differentiate between affinity, specificity, cellular action, and most important mechanism of action.Read more
Application Note Fragment Based Drug Discovery on Pioneer Systems Using Next Generation SPR Analysis
Fragment-based drug design (FBDD) has become an increasingly popular platform for the identification of lead candidates in drug discovery programs.Read more
Application Note OneStep Lead Characterization of High Affinity Biologic Interactions with Pioneer SPR Systems
Biopharmaceutical drug discovery and development have celebrated the approval of breakthrough treatments in diseases of inflammation, cancer and infectious disease in recent years.Read more
Application Note Biomolecular Binding Kinetics Assays on the Octet Platform
Direct measurement of biomolecular interactions plays an important role in biotherapeutic drug discovery and development. Label-Free analytical technologies such as the Octet® platform from Pall ForteBio provide a powerful means to obtain accurate information about rate of biomolecular complex formation and complex stability, key components of a drug-target interaction.Read more
Application Note Analysis of Fc-gamma Receptor-IgG Interactions on the Octet Platform
Fc gamma receptors (FcγRs) are membrane glycoproteins with affinity for the Fc region of immunoglobulin G (IgG). FcγRs expressed on the surface of immune effector cells play a key role in initiating Fc effector functions such as antibody-mediated cell-dependent cytotoxicity (ADCC)1, which is a major mechanism of action of therapeutic monoclonal antibodiesRead more
Application Note Octet Potency Assay: Development, Qualification and Validation Strategies
Kinetic analysis of biomolecular interactions is critical during drug discovery and development. The affinity of an interaction directly affects the dose required for a biopharmaceutical to be effective.Read more
Technical Guide Small Molecule Binding Kinetics
Small molecule kinetics can be measured on the Octet RED, Octet RED96 and Octet RED384 instruments. In a typical experiment, a biotinylated protein target is immobilized onto a high-capacity Super Streptavidin (SSA) biosensor surface, and this surface is exposed to a solution of the small molecule in a microplate well.Read more