Bachmann T T, Leca B, Vilatte F, Marty J L, Fournier D, Schmid R D
Institute for Technical Biochemistry, University of Stuttgart, Germany.
Biosens Bioelectron. 2000 Jun;15(3-4):193-201. doi: 10.1016/s0956-5663(00)00055-5.
Engineered variants of Drosophila melanogaster acetylcholinesterase (AChE) were used as biological receptors of AChE-multisensors for the simultaneous detection and discrimination of binary mixtures of cholinesterase-inhibiting insecticides. The system was based on a combination of amperometric multielectrode biosensors with chemometric data analysis of sensor outputs using artificial neural networks (ANN). The multisensors were fully manufactured by screen-printing, including enzyme immobilisation. Two types of multisensors were produced that consisted of four AChE variants each. The AChE mutants were selected in order to obtain high resolution, enhanced sensitivity and minimal assay time. This task was successfully achieved using multisensor I equipped with wild-type Drosophila AChE and mutants Y408F, F368L, and F368H. Each of the AChE variants was selected on the basis of displaying an individual sensitivity pattern towards the target analytes. For multisensor II, the inclusion of F368W, which had an extremely diminished paraoxon sensitivity, increased the sensor's capacity even further. Multisensors I and II were both used for inhibition analysis of binary paraoxon and carbofuran mixtures in a concentration range 0-5 microg/l, followed by data analysis using feed-forward ANN. The two analytes were determined with prediction errors of 0.4 microg/l for paraoxon and 0.5 microg/l for carbofuran. A complete biosensor assay and subsequent ANN evaluation was completed within 40 min. In addition, multisensor II was also investigated for analyte discrimination in real water samples. Finally, the properties of the multisensors were confirmed by simultaneous detection of binary organophosphate mixtures. Malaoxon and paraoxon in composite solutions of 0-5 microg/l were discriminated with predication errors of 0.9 and 1.6 microg/l, respectively.
果蝇乙酰胆碱酯酶（AChE）的工程变体被用作AChE多传感器的生物受体，用于同时检测和区分胆碱酯酶抑制性杀虫剂的二元混合物。该系统基于安培多电极生物传感器与使用人工神经网络（ANN）对传感器输出进行化学计量学数据分析的组合。多传感器完全通过丝网印刷制造，包括酶固定化。生产了两种类型的多传感器，每种由四种AChE变体组成。选择AChE突变体以获得高分辨率、增强的灵敏度和最短的分析时间。使用配备野生型果蝇AChE以及突变体Y408F、F368L和F368H的多传感器I成功完成了这项任务。每个AChE变体都是根据对目标分析物显示出独特的灵敏度模式而选择的。对于多传感器II，加入对氧磷敏感性极低的F368W进一步提高了传感器的能力。多传感器I和II都用于浓度范围为0 - 5微克/升的对氧磷和呋喃丹二元混合物的抑制分析，随后使用前馈ANN进行数据分析。两种分析物的测定预测误差分别为对氧磷0.4微克/升和呋喃丹0.5微克/升。完整的生物传感器分析及随后的ANN评估在40分钟内完成。此外，还研究了多传感器II对实际水样中分析物的区分能力。最后，通过同时检测二元有机磷混合物证实了多传感器的性能。在0 - 5微克/升的复合溶液中，对马拉硫磷和对氧磷进行了区分，预测误差分别为0.9微克/升和1.6微克/升。
