Mapping of Enzyme Kinetics on a Microfluidic Device

PLoS One. 2016 Apr 15;11(4):e0153437. doi: 10.1371/journal.pone.0153437. eCollection 2016.

Abstract

A microfluidic platform or "microfluidic mapper" is demonstrated, which in a single experiment performs 36 parallel biochemical reactions with 36 different combinations of two reagents in stepwise concentration gradients. The volume used in each individual reaction was 36 nl. With the microfluidic mapper, we obtained a 3D enzyme reaction plot of horseradish peroxidase (HRP) with Amplex Red (AR) and hydrogen peroxide (H2O2), for concentration ranges of 11.7 μM to 100.0 μM and 11.1 μM to 66.7 μM for AR and H2O2, respectively. This system and methodology could be used as a fast analytical tool to evaluate various chemical and biochemical reactions especially where two or more reagents interact with each other. The generation of dual concentration gradients in the present format has many advantages such as parallelization of reactions in a nanoliter-scale volume and the real-time monitoring of processes leading to quick concentration gradients. The microfluidic mapper could be applied to various problems in analytical chemistry such as revealing of binding kinetics, and optimization of reaction kinetics.

Publication types

  • Research Support, Non-U.S. Gov't
  • Validation Study

MeSH terms

  • Calibration
  • Equipment Design
  • Horseradish Peroxidase / chemistry
  • Horseradish Peroxidase / metabolism*
  • Hydrogen Peroxide / metabolism*
  • Imaging, Three-Dimensional
  • Kinetics
  • Lab-On-A-Chip Devices* / standards
  • Microfluidic Analytical Techniques* / instrumentation
  • Microfluidic Analytical Techniques* / methods
  • Microfluidic Analytical Techniques* / standards
  • Microscopy, Fluorescence
  • Time-Lapse Imaging

Substances

  • Hydrogen Peroxide
  • Horseradish Peroxidase

Grants and funding

This work was supported by the BE-Basic foundation (funded by the Ministry of Economic Affairs of the Netherlands, grant number: FES0905), a public private partnership of knowledge institutes, industry and academia, under the project nr. FS2.003. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.