Extracting kinetic rate constants from surface plasmon resonance array systems
ABSTRACT Surface plasmon resonance imaging systems, such as Flexchip from Biacore, are capable of monitoring hundreds of reaction spots simultaneously within a single flow cell. Interpreting the binding kinetics in a large-format flow cell presents a number of potential challenges, including accounting for mass transport effects and spot-to-spot sample depletion. We employed a combination of computer simulations and experimentation to characterize these effects across the spotted array and established that a simple two-compartment model may be used to accurately extract intrinsic rate constants from the array under mass transport-limited conditions. Using antibody systems, we demonstrate that the spot-to-spot variability in the binding kinetics was <9%. We also illustrate the advantage of globally fitting binding data from multiple spots within an array for a system that is mass transport limited.
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ABSTRACT: The simultaneous detection of multiple biological analytes is achieved for the first time using a single optical fiber based surface plasmon resonance (SPR) biosensor. This is achieved by collecting the plasmonic wave re-scattered by a rough metallic coating deposited onto two separate sections (sensing regions) of a single multimode optical fiber. The results obtained showed that two gastric cancer biomarkers (apolipoprotein E and clusterin) are detected in clinically relevant concentrations each on a separate sensing region, simultaneously, something that cannot be done in traditional fiber based SPR biosensors that read the transmitted or reflected light to monitor the position of the resonance. While this multiplexing demonstration has been performed with two different biomarkers, it paves the way for the multiplexed detection of a larger number of biomarkers using a simple fiber optic based SPR sensor for point of decision diagnostics.Sensors and Actuators B Chemical 07/2013; 5:454-458. DOI:10.1016/j.snb.2013.03.131 · 3.84 Impact Factor
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ABSTRACT: To explore the variability in biosensor studies, 150 participants from 20 countries were given the same protein samples and asked to determine kinetic rate constants for the interaction. We chose a protein system that was amenable to analysis using different biosensor platforms as well as by users of different expertise levels. The two proteins (a 50-kDa Fab and a 60-kDa glutathione S-transferase [GST] antigen) form a relatively high-affinity complex, so participants needed to optimize several experimental parameters, including ligand immobilization and regeneration conditions as well as analyte concentrations and injection/dissociation times. Although most participants collected binding responses that could be fit to yield kinetic parameters, the quality of a few data sets could have been improved by optimizing the assay design. Once these outliers were removed, the average reported affinity across the remaining panel of participants was 620 pM with a standard deviation of 980 pM. These results demonstrate that when this biosensor assay was designed and executed appropriately, the reported rate constants were consistent, and independent of which protein was immobilized and which biosensor was used.Analytical Biochemistry 12/2008; 386(2):194-216. DOI:10.1016/j.ab.2008.11.021 · 2.31 Impact Factor
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ABSTRACT: ForteBio's Octet optical biosensor harnesses biolayer interferometry to detect and quantify molecular interactions using disposable fiber-optic biosensors that address samples from an open shaking microplate without any microfluidics. We recruited a monoclonal antibody against a panel of peptides to compare the Octet directly with Biacore's well-established 3000 platform and Bio-Rad's recently launched ProteOn XPR36 array system, which use surface plasmon resonance (SPR) to detect the binding of one analyte over four surfaces and six analytes over six surfaces, respectively. A sink method was used to prevent analyte from rebinding the ligand-coated Octet tips and enabled us to extract accurate kinetic rate constants, as judged by their close agreement with those determined by SPR. Although the Octet is not sensitive enough to detect the binding of small molecules directly, it can access their affinities indirectly via solution competition experiments. We conducted similar experiments on the SPR instruments to validate these measurements. The Octet is emerging as a versatile complement to other more sophisticated biosensors, and the ProteOn provides high-quality data near the sensitivity of Biacore but in a more multiplexed format. Our results provide a benchmark for assessing the performance of the above-mentioned sensors.Analytical Biochemistry 07/2008; 377(2):209-17. DOI:10.1016/j.ab.2008.03.035 · 2.31 Impact Factor