The cluster glycoside effect.

Osteoprotegerin Is a Receptor for the Cytotoxic Ligand TRAIL

The past decade has seen researchers develop and apply novel technologies for biomolecular detection, at times approaching hard limits imposed by physics and chemistry. In nearly all sensors, the transport of target molecules to the sensor can play as critical a role as the chemical reaction itself in governing binding kinetics, and ultimately performance. Yet rarely does an analysis of the interplay between diffusion, convection and reaction motivate experimental design or interpretation. Here we develop a physically intuitive and practical understanding of analyte transport for researchers who develop and employ biosensors based on surface capture. We explore the qualitatively distinct behaviors that result, develop rules of thumb to quickly determine how a given system will behave, and derive order-of-magnitude estimates for fundamental quantities of interest, such as fluxes, collection rates and equilibration times. We pay particular attention to collection limits for micro- and nanoscale sensors, and highlight unexplained discrepancies between reported values and theoretical limits.

/pdf/making-it-stick-convection-reaction-and-diffusion-in-surface-2qlr7fj7la.pdf

Making it stick: convection, reaction and diffusion in surface-based biosensors.

The quality of optical biosensor data must be improved in order to characterize the mechanism and rate constants associated with molecular interactions. Many of the artifacts associated with binding data can be minimized or eliminated by designing the experiment properly, collecting data under optimum conditions and processing the data with reference surfaces. It is possible to globally fit high-quality biosensor data with simple bimolecular reaction models, which validates the technology as a biophysical tool for interaction analysis.

/pdf/improving-biosensor-analysis-588xhvziq4.pdf

Improving biosensor analysis.

We report here the evolution of ankyrin repeat (AR) proteins in vitro for specific, high-affinity target binding. Using a consensus design strategy, we generated combinatorial libraries of AR proteins of varying repeat numbers with diversified binding surfaces. Libraries of two and three repeats, flanked by 'capping repeats,' were used in ribosome-display selections against maltose binding protein (MBP) and two eukaryotic kinases. We rapidly enriched target-specific binders with affinities in the low nanomolar range and determined the crystal structure of one of the selected AR proteins in complex with MBP at 2.3 A resolution. The interaction relies on the randomized positions of the designed AR protein and is comparable to natural, heterodimeric protein-protein interactions. Thus, our AR protein libraries are valuable sources for binding molecules and, because of the very favorable biophysical properties of the designed AR proteins, an attractive alternative to antibody libraries.

High-affinity binders selected from designed ankyrin repeat protein libraries.

Surface plasmon resonance based biosensors are being used to define the kinetics of a wide variety of macromolecular interactions. As the popularity of this approach grows, experimental design and data analysis methods continue to evolve. These advances are making it possible to accurately define the assembly mechanisms and rate constants associated with macromolecular interactions.

Kinetic analysis of macromolecular interactions using surface plasmon resonance biosensors.

http://www.lmm.jussieu.fr/~lagree/TEXTES/BIACORE/583.pdf

Extending the Range of Rate Constants Available from BIACORE: Interpreting Mass Transport-Influenced Binding Data

Interpreting complex binding kinetics from optical biosensors: a comparison of analysis by linearization, the integrated rate equation, and numerical integration.

http://homes.nano.aau.dk/lg/biosensors2009_files/Myszka_Kinetic%20Analysis_BiophysChem64_127_1997.pdf

Thomas A. Morton

Papers

Kinetic analysis of macromolecular interactions using surface plasmon resonance biosensors.

Extending the Range of Rate Constants Available from BIACORE: Interpreting Mass Transport-Influenced Binding Data

Interpreting complex binding kinetics from optical biosensors: a comparison of analysis by linearization, the integrated rate equation, and numerical integration.

Kinetic analysis of a protein antigen-antibody interaction limited by mass transport on an optical biosensor.

CLAMP©: a biosensor kinetic data analysis program