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Related Experiment Video

Updated: Jun 28, 2026

Benchtop Immobilized Metal Affinity Chromatography, Reconstitution and Assay of a Polyhistidine Tagged Metalloenzyme for the Undergraduate Laboratory

Benchtop Immobilized Metal Affinity Chromatography, Reconstitution and Assay of a Polyhistidine Tagged Metalloenzyme for the Undergraduate Laboratory

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Enzyme-less amperometric biosensor for l-ascorbate using poly-l-histidine-copper complex as an alternative

Y Hasebe¹, T Akiyama, T Yagisawa

¹Department of Environmental Engineering, Faculty of Engineering, Saitama Institute of Technology, 1690, Fusaiji, Okabe, Saitama, 369-0293, Japan.

|October 31, 2008

Summary

A novel enzyme-less biosensor utilizing a poly-l-histidine(PLH)-copper(II) complex offers a stable and precise alternative for detecting l-ascorbate (AsA). This biocatalyst demonstrates comparable performance to traditional sensors and stability over extended use.

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A Rapid and Specific Microplate Assay for the Determination of Intra- and Extracellular Ascorbate in Cultured Cells

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Published on: April 11, 2014

Area of Science:

Biocatalysis
Biosensor Technology
Analytical Chemistry

Background:

Traditional l-ascorbate (AsA) sensors often rely on enzymes, which can be unstable.
Developing enzyme-less alternatives is crucial for robust biosensing applications.

Purpose of the Study:

To investigate the potential of a poly-l-histidine(PLH)-copper(II) complex as an enzyme-less biocatalyst for AsA detection.
To evaluate the performance, stability, and selectivity of this novel sensor system.

Main Methods:

Preparation of a PLH-copper(II) membrane by entrapping PLH in polyacrylamide gel and treating with CuCl(2).
Amperometric detection of AsA using the prepared membrane.
Assessment of sensor performance across a wide pH range and in the presence of potential interferents.

Main Results:

The PLH-Cu(II) complex functions as an effective enzyme-less biocatalyst for AsA detection.
The sensor exhibits precise determination of AsA (RSD < 3%) over a broad pH range (4-11).
Performance metrics are comparable to enzyme-based sensors, with excellent stability (over 3 months/300 samples) but limited selectivity against quinones.

Conclusions:

The PLH-Cu(II) complex presents a viable, stable, and enzyme-less alternative for AsA sensing.
The sensor demonstrates practical applicability in analyzing AsA in food and beverage samples.
Further optimization for selectivity may enhance its utility in complex matrices.