Elsevier

Biosensors and Bioelectronics

Volume 24, Issue 3, 15 November 2008, Pages 475-479
Biosensors and Bioelectronics

Short communication
Development of acetylcholinesterase biosensor based on CdTe quantum dots/gold nanoparticles modified chitosan microspheres interface

https://doi.org/10.1016/j.bios.2008.05.005Get rights and content

Abstract

In this paper, a novel acetylcholinesterase (AChE) biosensor was constructed by modifying glassy carbon electrode with CdTe quantum dots (QDs) and excellent conductive gold nanoparticles (GNPs) though chitosan microspheres to immobilize AChE. Since GNPs have shown widespread use particularly for constructing electrochemical biosensors through their high electron-transfer ability, the combined AChE exhibited high affinity to its substrate and thus a sensitive, fast and cheap method for determination of monocrotophos. The combination of CdTe QDs and GNPs promoted electron transfer and catalyzed the electro-oxidation of thiocholine, thus amplifying the detection sensitivity. This novel biosensing platform based on CdTe QDs–GNPs composite responded even more sensitively than that on CdTe QDs or GNPs alone because of the presence of synergistic effects in CdTe-GNPs film. The inhibition of monocrotophos was proportional to its concentration in two ranges, from 1 to 1000 ng mL−1 and from 2 to 15 μg mL−1, with a detection limit of 0.3 ng mL−1. The proposed biosensor showed good precision and reproducibility, acceptable stability and accuracy in garlic samples analysis.

Introduction

Recently, the applications based on nanoparticles (NPs) in electrochemistry have attracted wide interest due to their unique optical, electronic and mechanic properties (Wu et al., 2007a, Wu et al., 2007b). Various NPs such as gold (Zotti et al., 2008), silver (Chen et al., 2007) silica (Sun et al., 2006) and quantum dots (QDs) (Zhang et al., 2007) have been extensively used for ultrasensitive optical and electrochemical assays. With unique chemical and physical properties, gold nanoparticles (GNPs) have shown widespread use particularly for constructing electrochemical biosensors through their high electron-transfer ability between biomolecules and electrode surface (Kumar et al., 2007). Willner's group has extensively studied the electrochemical and optical applications of GNPs and the ability to promote the electron transfer (Willner et al., 2005).

Recently many uses of quantum dots (QDs) in biology were reported, such as cellular labeling (Voura et al., 2004, Chen and Gerion, 2004), in vivo tissue imaging (Gao et al., 2004), QDs assay labeling (Goldman et al., 2004, Tan et al., 2007). QDs-based electrochemical bioassay has also become a favorite topic because of inherent miniaturization, high sensitivity, low cost, and low power requirements. Its ability to promote the direct electron transfer between the biomolecules and electrode surfaces was also explored (Huang et al., 2005, Lu et al., 2005).

Chitosan-based materials have been extensively applied as immobilization matrices for preparation of biosensors (Ding et al., 2007, Tkac et al., 2007, Wu et al., 2007a, Wu et al., 2007b). Chitosan could be formulated as films, beads, microspheres and nanoparticles (Giunchedi et al., 1998). More importantly, chitosan micro/nanoparticles could be spontaneously formed through ionic gelation using tripolyphosphate as the precipitating agent, so that the use of organic solvents can be avoided during preparation and loading (Van der Lubben et al., 2001). And the chitosan microsphere/nanoparticles showed improved immobilization capacity for enzyme and good biocompatibility for preserving the activity of immobilized enzyme (Lin et al., 2007).

Acetylcholinesterase (AChE) is the most important target molecule of organophosphates (OP) compounds. In our previous work, an AChE biosensor based on silica sol–gel film assembling GNPs has been proposed. However, the conductivity of silica sol–gel material is not very good (Du et al., 2007a, Du et al., 2007b). In the present work, we constructed a novel interface with CdTe QDs and excellent conductive GNPs though chitosan microspheres to immobilize AChE, leading to a sensitive method for rapid determination of monocrotophos (Fig. 1). To the best of our knowledge, however, there has been no report focused on the synergy effect between CdTe QDs and GNPs to facilitate electron-transfer processes and the action of the immobilized AChE for OP determination. This new electrochemical system based on CdTe QDs–GNPs electrode responded even more sensitively than those modified by CdTe QDs or GNPs alone.

Section snippets

Reagents

AChE (Type C3389, 500 U mg−1 from electric eel), acetylthiocholine chloride (ATCl), N-hydroxysuccinimide (NHS), 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC) and sodium tripolyphosphate (TPP) were purchased from Sigma–Aldrich (St. Louis, USA) and used as received. UV spectra were recorded using a UV-2501 spectrophotometer (Shimadzu; Kyoto Japan). Monocrotophos and gold (III) chloride hydrate (3.9 g mL−1) were purchased from Treechem Co. (Shanghai, China).

Surface hydrophilicity

To gain better understanding of modification processes, the surface hydrophilicity was characterized by measuring their contact angles, as shown in Fig.S2 The CM/GCE displayed a small decrease of contact angle and thus a better hydrophilic surface than bare GCE. However, the obtained GNPs-CM/GCE led to an obvious decrease of contact angle, indicating a remarkable improvement of surface hydrophilicity due to unique chemical and physical properties of GNPs. Subsequent assembly of CdTe QDs on the

Conclusions

A simple enzyme biosensor based on immobilization of AChE on CdTe QDs–GNPs composite modified chitosan microspheres interface was developed for the amperometric determination of organophosphate pesticide. The formation of CdTe QDs–GNPs composite not only favored the interface enzymatic hydrolysis reaction to form electroactive substance, increasing the sensitivity and facilitating the amperometric response of the biosensor but also prevented enzyme molecule from leaking out of the electrode

Acknowledgments

The authors are gratefully acknowledging the financial support of the National Natural Science Foundation of China (Nos. 20705010, 20775064, 20735002, 20772038), the Research Fund for the Doctoral Program of Higher Education (No. 20070511015), the Program for Distinguish Young Scientist of Hubei Province (2007ABB017) and Program for Chenguang Young Scientist for Wuhan (200750731283).

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