Characterization of nanozyme kinetics for highly sensitive detection

Area of Science:

• Biochemistry
• Nanotechnology
• Analytical Chemistry

Background:

• Nanozymes are increasingly utilized as substitutes for natural enzymes like horseradish peroxidase (HRP).
• A significant gap exists in understanding the kinetic differences, specifically Michaelis-Menten constants (Km), between nanozymes and HRP.
• Over 60% of publications overlook these kinetic variations in analytical applications, potentially limiting nanozyme performance.

Purpose of the Study:

• To investigate the kinetic differences (Km) between peroxidase-mimicking nanozymes and HRP.
• To highlight the impact of these differences on analytical assay sensitivity.
• To demonstrate the necessity of optimizing substrate concentrations for nanozyme-based detection.

Main Methods:

• Conducted a comprehensive literature survey of 261 publications on nanozymes.
• Characterized the kinetic parameters (Km) of a gold-platinum (Au@Pt) nanozyme using TMB and H2O2 substrates.
• Applied the Au@Pt nanozyme as a label in immunoassays to evaluate detection limits under varying substrate concentrations.

Main Results:

• Identified significant differences in Km values between nanozymes and HRP across numerous studies.
• Found that the Au@Pt nanozyme exhibited a Km for TMB comparable to HRP but a Km for H2O2 three orders of magnitude higher.
• Demonstrated that non-optimized substrate concentrations resulted in a 30-fold increase in the limit of detection (LOD) compared to optimized conditions.

Conclusions:

• Measuring nanozymes' kinetic parameters is essential for accurate analytical development.
• Adjusting substrate concentrations based on determined kinetic parameters significantly enhances detection sensitivity.
• Optimized kinetic analysis is critical for unlocking the full potential of nanozymes in sensitive detection assays.