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Anal Biochem. 2010 Oct 1;405(1):1-10. doi: 10.1016/j.ab.2010.06.009. Epub 2010 Jun 8.

Comparison of label-free biosensing in microplate, microfluidic, and spot-based affinity capture assays.

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Department of Electrical and Computer Engineering, University of Illinois at Urbana-Champaign, Micro and Nanotechnology Laboratory, Urbana, IL 61801, USA.


Using both experimental assays and fluid-dynamic finite element simulation models, we directly compared the achievable performance limits of four distinct assay configurations for label-free detection of an analyte from a test sample on a biosensor surface. The assay configurations studied in this work included a biosensor incorporated into the bottom surface of a microplate well and a microfluidic channel. For each configuration, we compared assay performance for the scenario in which the entire bottom surface of the fluid-handling vessel is coated with capture ligands with assay performance for the scenario in which the capture ligands are applied in the form of localized spots. As a model system, we used detection of the protein biomarker tumor necrosis factor-alpha (TNF-alpha) using immobilized TNF-alpha capture antibody. Results show that the microfluidic assay format dramatically reduces the time required to establish a stable equilibrium. Spot-based assays are advantageous for microplate-based detection for reducing the time required for equilibrium sensor response. The results derived are generally applicable to any label-free biosensor technology and any ligand-analyte system with adjustable variables that include sensor mass density sensitivity, analyte-ligand adsorption/desorption rate constants, immobilized ligand density, flow channel geometry, flow rate, and spot size.

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