An on-chip fluorogenic enzyme assay using a multilayer microchip interconnected with a nanocapillary array membrane

Maojun Gong, Bo Young Kim, Bruce R. Flachsbart, Mark A. Shannon, Paul W. Bohn, Jonathan V. Sweedler

Research output: Contribution to journalArticlepeer-review

Abstract

Microfluidic devices allow manipulation of reagents and fluids in a semi-automated fashion, ideal for performing multiple measurements or conditioning various reagents. Here, an enzyme assay has been performed in a multilayer poly(methyl methacrylate)-based microfluidic device, where the layers are fluidically connected via a nanocapillary array membrane serving as an effective injector and valve. As a model system, β-glucuronidase from Escherichia coli and fluorescein di(β-D-glucuronide) are used for the assay; offline mixing and online incubation of substrate and enzyme allow determination of the initial hydrolysis rates of the substrate under catalysis by β-glucuronidase. The Michaelis constant Km was determined to be ∼ 4.0 μM for the enzyme of 83 units/mL at ambient temperature. The 50% inhibitory concentration IC50 of D-saccharic acid-1,4-Iactone to 167 units/mL was estimated to be 3.0 μtM. These results demonstrate added functionality for a poly(methyl methacrylate)-based nanocapillary array membrane-containing microfluidic device for following enzyme reaction kinetics.

Original languageEnglish (US)
Article number4529140
Pages (from-to)601-607
Number of pages7
JournalIEEE Sensors Journal
Volume8
Issue number5
DOIs
StatePublished - May 2008

Keywords

  • Enzyme kinetics
  • Microcontact printing
  • Microfluidics
  • Nanocapillary array membranes
  • Nanofluidics
  • Poly(methyl methacrylate)

ASJC Scopus subject areas

  • Engineering(all)
  • Electrical and Electronic Engineering

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