Design and characterization of immobilized enzymes in microfluidic systems

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Abstract

Herein we report the fabrication, characterization, and use of total analytical microsystems containing surface-immobilized enzymes. Streptavidin-conjugated alkaline phosphatase was linked to biotinylated phospholipid bilayers coated inside poly(dimethylsiloxane) microchannels and borosilicate microcapillary tubes. Rapid determination of enzyme kinetics at many different substrate concentrations was made possible by carrying out laminar flowcontrolled dilution on-chip. This allowed Lineweaver-Burk analysis to be performed from a single experiment with all the data collected simultaneously. The results revealed an enzyme turnover number of 51.1 ± 3.2 s-1 for this heterogeneous system. Furthermore, the same enzyme immobilization strategy was extended to demonstrate that multiple chemical reactions could be performed in sequence by immobilizing various enzymes in series. Specifically, the presence of glucose was detected by two coupled steps employing immobilized avidinD-conjugated glucose oxidase and streptavidin-conjugated horseradish peroxidase.

Original languageEnglish (US)
Pages (from-to)379-385
Number of pages7
JournalAnalytical chemistry
Volume74
Issue number2
DOIs
StatePublished - Jan 15 2002

All Science Journal Classification (ASJC) codes

  • Analytical Chemistry

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