Immobilization of multi-enzyme microreactors inside microfluidic devices

Won Gun Koh, Michael Pishko

Research output: Contribution to journalArticlepeer-review

98 Citations (Scopus)


A simple method to fabricate enzyme-containing microscopic hydrogel structures in microfluidic devices for the potential use in micro total analysis systems (μ-TAS) is described. Poly(ethylene glycol)-based hydrogel microstructures were prepared inside microchannels by photolithography and enzymes conjugated to a pH sensitive fluorophore (SNAFL-1) were incorporated into these hydrogel microstructures. Because of the ratiometric pH-dependent nature of SNAFL fluorescence, hydrogel microstructures exhibited a different emission intensity ratio with pH and this intensity ratio changed almost linearly between pH 7 and 12. When alkaline phosphatase-containing microreactors were exposed to p-nitrophenylphosphate (pNPP) as a substrate, phosphoric acid was produced inside the microstructure by enzymatic-catalyzed hydrolysis of the substrate and subsequently decreased the microenvironment pH. Because of the relatively rapid mass transport of analyte through the hydrogel, enzyme-catalyzed reaction was easily detected by change in emission intensity ratio before and after exposure to substrates. Enzyme-catalyzed reactions were quite fast and reached 90% of maximum value within 10 min. Data were analyzed using a modified Michaelis-Menten equation and apparent Michaelis constants could be obtained. This system was also successfully applied to urea hydrolysis by urease.

Original languageEnglish
Pages (from-to)335-342
Number of pages8
JournalSensors and Actuators, B: Chemical
Issue number1 SPEC. ISS.
Publication statusPublished - 2005 Apr 29

Bibliographical note

Funding Information:
The work described in this paper was supported by a grant from the National Aeronautics and Space Administration (NASA, NAG 91277).

All Science Journal Classification (ASJC) codes

  • Electronic, Optical and Magnetic Materials
  • Instrumentation
  • Condensed Matter Physics
  • Surfaces, Coatings and Films
  • Metals and Alloys
  • Electrical and Electronic Engineering
  • Materials Chemistry


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