Amperometric PMMA-microchip with integrated gold working electrode for enzyme assays

Area of Science:

• Analytical Chemistry
• Biochemistry
• Microfluidics

Background:

• Alkaline phosphatase (AP) enzymatic assays are crucial in diagnostics and research.
• Existing methods may require complex instrumentation or longer analysis times.
• Microchip electrophoresis offers a miniaturized platform for rapid separations.

Purpose of the Study:

• To develop and validate a microchip capillary electrophoresis (CE) system for alkaline phosphatase (AP) enzymatic assays.
• To evaluate the performance of a gold film in-channel detector integrated with a poly(methyl methacrylate) (PMMA) CE microchip.
• To assess the feasibility of rapid, reagent-inclusive enzymatic assays.

Main Methods:

• Utilized a poly(methyl methacrylate) (PMMA) CE microchip with an integrated gold film in-channel detector.
• Employed Tris-borate or Tris-Gly buffer solutions (pH 9.0, 50 mmol L(-1)) as running buffers.
• Investigated the detection of AP products (alpha-naphthol, p-nitrophenol, ascorbic acid) and assessed assay reproducibility and concentration dependence.

Main Results:

• Achieved reproducible signals for common AP products with a relative standard deviation (RSD) of 4.4% for alpha-naphthol.
• Demonstrated a concentration-dependent response, with a linear relationship observed for ascorbic acid between 5 and 20 mmol L(-1).
• Showcased the potential for rapid enzymatic assays (under 3 minutes) by incorporating reagents into the running buffer.

Conclusions:

• The gold film in-channel detector coupled with a PMMA CE microchip is a viable platform for AP enzymatic assays.
• The system provides reliable and sensitive detection of AP products.
• The integration of reagents into the running buffer enables highly efficient and rapid enzymatic assays.