One-step enzyme kinetics measurement in 3D printed microfluidics devices based on a high-performance single vibrating...

Area of Science:

• Biochemistry
• Microfluidics
• Enzyme kinetics

Background:

• Enzyme kinetics are crucial for understanding biological processes and applications in biosensing and industry.
• Conventional methods are laborious, consume excess reagents, and struggle with fast kinetics.

Purpose of the Study:

• To develop a simplified, one-step assay for measuring enzyme kinetics.
• To leverage 3D-printed microfluidics and advanced mixing for efficient kinetic analysis.

Main Methods:

• Utilized a 3D-printed microfluidic device with a vibrating sharp-tip mixer for rapid fluid mixing (∼300 μm, 3 ms).
• Performed enzyme kinetic experiments by adjusting reagent flow rates in a single run.
• Measured Michaelis-Menten kinetics for horseradish peroxidase (HRP) using hydrogen peroxide and amplex red.

Main Results:

• Achieved efficient mixing across a wide flow rate range (1.5–750 μL/min).
• Obtained Michaelis constant values consistent with literature data.
• Demonstrated a simplified workflow with reduced sample and reagent needs.

Conclusions:

• The developed one-step assay significantly simplifies enzyme kinetics measurement.
• The method offers high temporal resolution, low power consumption, and potential for optimizing biosensing and biochemical studies.