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-α) using immobilized TNF-α 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.

Original languageEnglish (US)
Pages (from-to)1-10
Number of pages10
JournalAnalytical Biochemistry
Issue number1
StatePublished - Oct 2010


  • Capture affinity assay
  • Label-free biosensor
  • Microfluidic
  • Microplate
  • Photonic crystal
  • Spot-based assay

ASJC Scopus subject areas

  • Biochemistry
  • Biophysics
  • Molecular Biology
  • Cell Biology


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