Assay design

Rich, Rebecca L. et al Biosensor-based fragment screening using FastStep injections. Analytical.Biochemistry 407: 270-277; (2010). Goto reference

The development a novel analyte injection method is described for the SensiQ Pioneer surface plasmon resonance-based biosensor referred to as "FastStep." By merging buffer and sample streams immediately prior to the reaction flow cells, the instrument is capable of automatically generating a two- or threefold dilution series (of seven or five concentrations, respectively) from a single analyte sample. Using sucrose injections, it is demonstrated that the production of each concentration within the step gradient is highly reproducible.

De Crescenzo, Gregory et al Online optimization of surface plasmon resonance-based biosensor experiments for improved throughput and confidence. Journal of Molecular Recognition 21: 256-266; (2008). Goto reference

The emergence of surface plasmon resonance-based optical biosensors has facilitated the identification of kinetic parameters for various macromolecular interactions. Normally, these parameters are determined from experiments with arbitrarily chosen periods of macromolecule and buffer injections, and varying macromolecule concentrations. Since the choice of these variables is arbitrary, such experiments may not provide the required confidence in identified kinetic parameters expressed in terms of standard errors. It is shown using multiple experimental and simulated data that the desired confidence can be reached with much shorter experiments than those generally performed by biosensor users.

Karlsson, R. et al Analyzing a kinetic titration series using affinity biosensors. Analytical Biochemistry 349: 136-147; (2006). Goto reference

The classical method of measuring binding constants with affinity-based biosensors involves testing several analyte concentrations over the same ligand surface and regenerating the surface between binding cycles. Here an alternative approach to collecting kinetic binding data is described, which is called "kinetic titration." This method involves sequentially injecting an analyte concentration series without any regeneration steps. In addition, kinetic titrations can be more efficient than the conventional data collection method and allow us to fully characterize analyte binding to ligand surfaces that are difficult to regenerate.

Myszka, D. G. Improving biosensor analysis. J.Mol.Recognit. 12: 279-284; (1999).

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.

Beattie, J. et al Molecular interactions in the insulin-like growth factor (IGF) axis: a surface plasmon resonance (SPR) based biosensor study. Mol.Cell Biochem. : (2007).

A comprehensive analysis of the interaction between insulin-like growth factor and six IGF binding proteins with detailed experimental analysis of the experimental design and solutions for encountered problems.

Szolar, O. H. J. et al Qualification and application of a surface plasmon resonance-based assay for monitoring potential HAHA responses induced after passive administration of a humanized anti Lewis-Y antibody. J.Pharm.Biomed.Anal. 41: 1347-1353; (2006).

Short publication with an outline for the qualification of a SPR-based concentration assay.

Giannetti, Anthony M. et al Surface Plasmon Resonance Based Assay for the Detection and Characterization of Promiscuous Inhibitors. Journal of Medicinal Chemistry 51: 574-580; (2008). Goto reference

Publication how do identify and deal with promiscuous interactants. This article outlines a systematic approach to identify and classify promiscuous binders. In addition, it presents a number of datasets from poorly behaved to well behaved.

Schasfoort, Richard B. M. et al Method for estimating the single molecular affinity. Analytical Biochemistry 421: 794-796; (2012). Goto reference

Affinity constants can be determined by methods that apply immobilized ligands such as immunoassays and label-free biosensor technologies. This article outlines a new surface plasmon resonance array imaging method that yields affinity constants that can be considered as the best estimate of the affinity constant for single biomolecular interactions. Calculated rate (kd and ka) and dissociation equilibrium (KD) constants for various ligand densities and analyte concentrations are extrapolated to the KD at the zero response level (KDR0).

Bedford, Erin E. et al Surface Plasmon Resonance Biosensors Incorporating Gold Nanoparticles. Macromolecular Bioscience 12: 724-739; (2012). Goto reference

This article describes the incorporation of gold nanoparticles in the setup of the experiment. Recent methods to enhance SPR detection capabilities using gold nanoparticles are reviewed, as well as device fabrication and the results of incorporation. SPR detection is a highly versatile method for the detection of biomolecules and, with the incorporation of AuNPs, shows promise in extending it to a number of new applications.