The development of antibodies for diagnostics or therapeutics requires comprehensive characterization of affinity, specificity, and mechanism of action (doi.org/10.1016/j.xpro.2021.100836). Affinity measurements screen different isolates of an antibody to identify those that are most effective at binding antigen. Also, measurements of nonspecific binding to compounds that are structurally related to the antigen of interest can help establish the likelihood of cross reactivity with other molecules that accompany the antigen.
Biolayer interferometry (BLI) is a label free technique that measures the interference pattern of white light reflected from the surface of a biosensor, which indicates the presence of biomolecular interactions. The binding between a ligand immobilized on the biosensor tip and an analyte in solution produces an increase in optical thickness at the biosensor tip, resulting in a wavelength shift proportional to the extent of binding (Azmiri and Lee, 2015; Mechaly et al., 2016). The sensor tips collect readings in real time, while immersed in the analyte solution, without the need for continuous flow fluidics (Yang et al., 2017).
BLI technology is widely used for quantitation of antibodies, which is fundamental to biological research and production processes. The technology is also extensively used for kinetics measurements of antibodies and small molecules to assess the strength and speed of binding of an antibody to a target. This is possible due to BLI’s robustness to complex matrices, speed, accuracy, and ease of use.
Biosensors with different ligands that bind antibodies, proteins and small molecules such as streptavidin, human Fc, mouse Fc, protein A, anti-his, and Ni-NTA are routinely used. However, many of the first generation BLI biosensors have limited dynamic range, poor small molecule binding, and higher costs due to single use operation. The wider adoption of this simple and yet powerful technology was also limited by relatively expensive instrumentation and complex software.
This e-Book explores advances in biosensor technology and instrumentation that can significantly accelerate antibody discovery through efficient quantitation, kinetics, and epitope binning.