A novel acetylcholinesterase (AChE)/choline oxidase (ChOx) bienzyme amperometric acetylcholine biosensor based on gold nanoparticles (AuNPs) and multi-walled carbon nanotubes (MWCNTs) has been successfully developed by self-assembly process in combination of sol–gel technique. A thiolated aqueous silica sol containing MWCNTs and ChOx was first dropped on the surface of a cleaned Pt electrode, and then AuNPs were assembled with the thiolated sol–gel network. Finally, the alternate deposition of poly (diallyldimethylammonium chloride) (PDDA) and AChE was repeated to assemble different layers of PDDA-AChE on the electrode for optimizing AChE loading. Among the resulting biosensors, the biosensor based on two layers of PDDA-AChE multilayer films showed the best performance. It exhibited a wide linear range, high sensitivity and fast amperometric response, which were 0.005–0.4 mM, 3.395 μA/mM, and within 15 s, respectively. The biosensor showed long-term stability and acceptable reproducibility. More importantly, this study could provide a simple and effective multienzyme immobilization platform for meeting the demand of the effective immobilization enzyme on the electrode surface.Highlights► A novel bienzyme amperometric acetylcholine biosensor based on gold nanoparticles and carbon nanotubes was developed by self-assembly process in combination of sol–gel technique. ► The biosensor was demonstrated as a high sensitivity and long-term stability tool for the analysis of acetylcholine. ► The proposed design can provide a simple and effective platform for constructing the multienzyme-based biosensor.

Plant pathogenic bacteria spread all over the world, causing a great deal of economic loss. This study has developed a novel method for rapid detection of plant pathogenic bacteria by electrochemiluminescence polymerase chain reaction (ECL-PCR) using two universal probes, a biotin-probe and a Ru(bpy)32+ (TBR)-probe. Biotin-probe sequence is complementary to the universal sequence of anti-sense primer, and TBR-probe sequence is the same as the universal sequence of sense primer. So the amplified PCR products can hybridize with TBR-probe and biotin-probe. After hybridizing, PCR products are captured by streptavidin coated magnetic bead through the biotin–streptavidin linkage, and then TBR reacts with tripropylamine to emit light for detection of plant pathogenic bacteria. This proposed method is applied to detect Fusarium oxysporum f. sp. Cubense and Xanthomonas oryzae pv. Oryzae. The results show that the method can successfully identify the plant pathogenic bacteria in the infected samples and these results are consistent with the results of gel electrophoresis. Importantly, this study can be used as an illustration for detecting various plant pathogenic bacteria and provides a feasible approach on developing ECL sensors to meet the demand of rapid detection of pathogenic bacteria, fungi, and viruses.