Saxitoxin (STX) is one of the paralytic shellfish poisons (PSP) that endanger people’s health. It is necessary to develop methods for the on-site rapid detection for STX in order to prevent safety accidents. An enzyme-linked immunosorbent assay (ELISA) is timesaving and effective, but it is not suitable for large-scale in-field tests due to the expensiveness of commercial ELISA kits and the bulkiness of a microtiter plate reader (MTPR). In this study, a portable smartphone-based colorimetric analyzer (SBCA) with a cost-effictive enhanced gold nanoparticle-based ELISA (EGNB-ELISA) was proposed for STX detection. In a bicinchoninic acid (BCA) protein assay (R2 = 0.9939) and a glucose assay (R2 = 0.9937), SBCA was shown to be in good agreement with MTPR. EGNB-ELISA had a 12.5-fold lower detection limit (0.4 ng/mL) and a lower detection range (1 - 50 ng/mL, Y = 0.4037X + 0.3564, R2 = 0.9797) than the classical ELISA. The recovery rate ranged over 89.1 - 112.2%. The whole detection system, combining both homemade SBCA and ENGB-ELISA, is expected to satisfy the needs of on-site STX sample tests to guarantee seafood safety.

A method combining a kit with the Bionic e-Eye for rapid and portable on site detection of diarrhetic shellfish poisoning.

Polylactic acid (PLA) and polycaprolactone (PCL) were selected as materials to fabricate tissue engineering scaffolds by three-dimensional (3D) printing and electrospinning ,which were used to culture cardiomyocytes of neonatal rats .Then the scaffolds with cardiomyocytes were coupled with microelectrode array (MEA) to form a 3D cell-based biosensor ,which was used to detect the extracellular field potential (EFP) of cardiomyocytes .The experimental results demonstrated that cardiomyocytes adhered and grew well in scaffolds ,and could drive fibers to produce combined beating due to the excitation-contraction coupling .After 48 hours ,the beating rate of cardiomyocytes in the scaffolds tended to be stable .The detecting results demonstrated that scaffolds and MEA were coupled well to be a 3D cell-based biosensor system ,which could detect the EFP of cardiomyocytes in scaffolds with stable and high-SNR signals .The EFP amplitude and firing rate were both similar to the signals recorded from traditional two-dimensional (2D) culturing method.

In order to solve the problems of low precision and single parameter in the traditional in-vitro rapid diag-nostic system,a high precision and multi-parameter rapid in-vitro diagnostic system is designed based on the mi-crofluidic and micro-nano sensing technology. The paper mainly includes the introduction and presentation of the following aspects:the microfluidic chip design,the micro-nano sensing preparation and processing technology,the multidimensional anti-interference correction algorithm based on microfluidic multi-channel reference technique,the weak biosensing signal extraction and acquisition,and the multi-parameter in-vitro rapid diagnostic analysis system design. The system has the advantages of high detection accuracy,good anti-jamming performance,and multi-param-eter integration. The basic performance of the multi-parameter rapid detection system is tested using systematic test and clinical blood. The experimental results show that the multi-parameter in-vitro diagnostic rapid detection system can be a comprehensive detection of blood glucose,four items of blood lipids,uric acid,blood ketone and many other indicators,with fast accurate and multi-parameter measurement,which may provide a practical technology platform for the detection of portable biochemical indicators of basic medical institutions.

Okadaic acid (OA) and saxitoxin (STX) are typical toxins of diarrhetic shellfish poisoning (DSP) and paralytic shellfish poisoning (PSP), respectively, which are highly toxic marine toxins threatening human health and environmental safety. OA is a potent inhibitor of serine/threonine protein phosphatases that can cause cellular death, while STX is an inhibitor of sodium channel that can lead to neurological damage. In this work, a dual functional cardiomyocyte-based biosensor was proposed to detect DSP and PSP toxins by monitoring the viability and electrophysiology of cardiomyocytes. The results showed that the viability of cardiomyocytes was sensitive to the OA and STX, resulting in significant changes of the electrophysiological properties, including amplitude, firing rate and duration of the extracellular field potential (EFP). The detection limits of the hybrid-biosensor are as low as 7.16 ng/mL for OA and 5.19 ng/mL for STX. In summary, all of the results indicate that the dual functional cardiomyocyte-based hybrid-biosensor will be a promising and utility tool for shellfish toxin detection.

A microplate method provides an efficient way to use modern detection technology. However, there are some difficulties concerning on-site detection, such as being non-portable and time-consuming. In this work, a novel portable microplate analyzer with a thermostatic chamber based on a smartphone was designed for rapid on-site detection. An analyzer with a wide-angle lens and an optical filter provides a proper environment for the microplate. A smartphone app-iPlate Monitor was used for RGB analyze of image. After a consistency experiment with a microtiter plate reader (MTPR), the normalized calibration curves were y = 0.7276x + 0.0243 (R2 = 0.9906) and y = 0.3207x + 0.0094 (R2 = 0.9917) with a BCA protein kit as well as y = 0.182x + 0.0134 (R2 = 0.994) and y = 0.0674x + 0.0003 (R2 = 0.9988) with a glucose kit. The times for obtaining the detection requirement were 15 and 10 min for the BCA protein kit and the glucose kit at 37°C; in contrast, it required more than 30 and 20 min at ambient temperature. Meanwhile, it also showed good repeatability for detections.

kadaic acid (OA), as a diarrheic shellfish poisoning toxin, had wide distribution and frequent occurrence. Therefore, low-cost, high-throughput, wide-range and portable detection of OA was in high demand for food safety and environmental monitoring. In this study, a novel and portable smartphone-based system using cell viability biosensor (CVBS) was developed for label-free, non-invasive and long-term monitoring of cell viability. The variation of cell viability reflected the changes of cell morphology, cell count and cell proliferation indirectly. And this system applied the combination of image analysis and cell counting kit-8 assay (CCK-8) to monitor the reflection. The biosensing system chose HepG2 cells as sensing elements to build CVBS and used it in OA detection. Results showed this system could synchronously detect OA in 96 channels. And this biosensor presented a good performance to various OA concentrations, with a wide linear detection range (10-800 mu g/L). Moreover, the point-in-time having best detection performance could be located by the traversal algorithm in the monitoring duration. Thus, this cell-based biosensor system provided a convenient and efficient approach in seafood safety testing such as OA screening. (C) 2017 Published by Elsevier B.V.

Saxitoxin( STX) are common marine toxins which can affect human health through food chain by accumu-lating in shellfishes. It is urgently required to have a convenient method for on-site detection of marine toxins to a-void the poisoning incidents that have occurred frequently. In this work,a handheld analysis meter system coopera-ting with competitive immunoassay strips is designed for rapid detection of STX. The smartphone is used as light de-tector,for image acquisition and data processing by a specific application. The 3D-printed portable accessory of sm-artphone is fixed for test strip. The method is simple and fast,with a detection limit of STX of 5.2 ng/mL,respec-tively. It can provide quantitative analysis of STX in the range of 5.2 ng/mL~100 ng/mL. Based on the smartphone system and biochemical analysis,it will be promising tool for on-site rapid detection for paralytic shellfish poisoning.

A novel bioanalysis method for sensitive, rapid and field-based detection of okadaic acid (OA) is in great demand for environmental monitoring and food security. In this study, we reported a biosensing system consisting of a smartphone-based system and cell viability biosensor (CVB) for OA spot fast detection. CVB was constructed by combing living cells and cell counting kit (CCK-8). Thereinto, living cells could be used as reagent without adherent culture on the device surface, and adherent cells and suspension cells were both applicable for cell type selection. The smartphone-based system- bionic electronic eye (Bionic e-Eye) was composed of portable illumination provider and smartphone. The homemade application program (App) - iPlate Monitor integrated the software functions of real-time image acquisition and analysis, data display and storage and instant data sharing. In the measurement, the system was demonstrated to detect OA at concentration as low as 3.4083 mu g/L and 13.4456 mu g/L by using HepG2 and THP-1 cells, respectively. In addition, this system distinguished OA versus different marine toxins such as gonyautoxin2&3 (GTX2&3) and brevetoxin-2 (PbTx-2) in high specificity. Consequently, the smartphone-based biosensing platform provided a convenient, low-cost, easy-to-use and efficient approach for spot rapid detection of marine toxins such as OA. (C) 2017 Elsevier B.V. All rights reserved.

Okadaic acid (OA) is a representative diarrhetic shellfish poisoning toxin which is highly toxic and carcinogenic to human, and rich in polluted shellfish. In this work, a novel and high sensitive fluorescence immunosensor based on magnetic beads (MBs) and quantum dots (QDs) had been developed for OA detection. Carboxylic acid modified MBs were used as supporter to immobilize the OA-BSA, competed with the OA in the sample solution to bind with the anti-OA monoclonal antibody (OA-MAb). The CdTe QDs (EX600 nm) labeled IgG was served as secondary antibody to perform fluorescence detection. A portable flow cytometry, Moxi Flow was applied for OA on-site quantification. The results showed that the OA concentration was inversely proportional to the QDs fluorescence intensity. The limit of detection (LOD) was 0.05 μg/L with a linear range of 0.2–20 μg/L for OA detection, which was far lower than traditional id-ELISA strategy. Moreover, OA detection for the real sample could be completed within 1 hour. The matrix effect and the recovery rate were also assessed during real sample measurement, showing a high recovery.

In order to achieve the requirement of diarrhetic shellfish poison(DSP)in￣suit detection,we design a high throughput pretreatment system and a mobile terminal based detection system. Our system can make fast and accurate DSP detection by analyzing image of chromogenic reaction come from ELISA method. The rate of recovery by our pretreatment system and by hand are 89% and 93% respectively,it means that our pretreatment system can meet the follow￣up detection requirement. Detect the prevented samples by our detection system, the standard deviation is approximately 0.13 and average rate of recovery is 89.5%,it can satisfy the accuracy and repeatability of detection. In the detection of actual samples,comparing our results with the results come from microplate reader,it shows that our pretreatment and detection system can do a fast and accurate in￣suit work and provide a new method for the detection.

为了构建高度稳定性和一致性的心肌细胞电位传感器,从微电极阵列表面亲水性和细胞培养密度两方面对心肌细胞和微电极阵列(MEA)的耦合性进行研究.通过对MEA表面进行高分子蛋白明胶的修饰来提高MEA表面的亲水性,并重点分析不同细胞密度下构建的心肌细胞电位传感器的胞外场电位信号(EFP)信号特征.研究结果表明:心肌细胞按优化密度12万/cm2培养在经明胶修饰的MEA表面上,可促使心肌细胞和MEA形成高度紧密的耦合.在此条件下构建的心肌细胞传感器,能稳定输出一致性良好的EFP信号,电位幅值可达到约1.2mV,发放频率可达到约180次/min,信号稳定期可维持3～4 d.通过选择2种典型的工具药物异丙肾上腺素和利多卡因对优化后的心肌细胞电位传感器进行分析性能的测试,实验结果表明:20 μM的异丙肾上腺素和利多卡因分别大幅度增强和抑制了电位幅值和发放频率,结果与文献报导的结果相一致.该心肌细胞传感器对2种测试药物作出了快速而灵敏的响应,有望成为药物检测和分析的有效平台.

Based onEnzyme-linked immunosorbent assay(ELISA),test kit,namely,enzyme sensor and the corre-sponding instrument was prepared,Bionic e-Eye for rapid,effective and quantitative detection of okadaic acid(OA) on-site. The detection limit of the kit is 0.19μg/L and the range is among 0.2μg/L and 5μg/L. The recovery rate is among 75%and 125%and the accuracy is within±15%. The performance of Bionic e-Eye is consistent with micro-plate reader and high performance liquid chromatography(HPLC). In addition,without artificial calculation,the instrument can show the results directly on the screen,which is very suitable for rapid detection,and will be a new technology for rapid and quantitative detection of OA on site.

In this study,the feasibility of utilizing male mouse germ cells in testis and electrical cell-substrate imped?ance sensor(ECIS)was explored to build a cell-based bitter biosensor,which can sensitively and specifically respond to bitter compounds. Male mouse germ cells express lots of bitter receptor T2Rs(G protein coupled receptors)which can sensitively and specifically respond to bitter compounds,when active by ligands will affect cell morphology in a specific manner. The best cell density was explored;cell-impedance response profiles of germ cells to two bitter com?pounds were investigated by analyzing the response intensity under various concentrations,The linear detection range of PTC is 10μmol/L~200μmol/L,the detection limit is 4μmol/L;the linear detection range of quinine is 62.5μmol/L~1000μmol/L,the detection limit is 4μmol/L; Furthermore,impedance responses to five basic tastes were exam?ined to evaluate the specificity of cell-based bitter biosensor in bitter detection. The results revealed that this hybrid bitter biosensor could respond to those bitter compounds in a dose-dependent manner. And it could respond to bitter compounds specifically. This biosensor may provide a promising approach for detecting various bitter compounds.

With the rapid development of modern industry,the threat from marine toxins to human health has be?come increasingly serious,especially the paralytic biological poison. This paper presents a new method of cell-based potential sensor to detect shellfish toxicity. The cardiomyocyte combined with microelectrode array(MEA)by microelectronic processing technique which construct a novel cardiomyocyte potential sensor. The sensor can not on?ly real-time monitoring,but also could output good extracellular field potential signal with stability and consistency, which cardiomyocytes seeded on the MEA high-closely. By testing saxitoxin(STX)and analyzing the characteristics of signal parameters,the experimental results illustrate the STX have obvious inhibitory effect for electrical activity of cardiomyocyte. And the peak potential amplitude and frequency have obvious inhibition of concentration depen?dence,and presented the different concentration sensitivity. The LOD(limit of detection)of conventional standard laboratory toxicity detection on mouse bioassay method is 40μg/100 g,while the LOD and detection range of cardio?myocyte potential sensor is 1.004 ng/mL and 25 nmol/L~1 600 nmol/L,respectively. By comparative experiments, this method has great LOD and the detection is relatively simple and easy to fit standardization. With further im?provement,the method has good prospects for development on detection rapidly toxicity of marine toxins.

Based on the inhibition of okadaic acid (OA) on protein phosphatase 2A (PP2A), we developed disposable poly-o-aminophenol-carbon nanotubes (PoAP-CNTs) modified screen print electrode (SPE)-based electrochemical enzyme sensor for OA detection. Electropolymerized PoAP-CNTs composite film was coated on electrode for the immobilization of PP2A. Owing to the excellent physicochemical characteristic of PoAP-CNTs film, the performance of enzyme sensor was enhanced. Under the optimal conditions, the biosensor provides a larger linear range, and lower detection limit for the analysis of OA compared with colorimetric PP2A inhibition assay and PP2A/SPE. The spiked OA shellfish samples were applied to verify the efficiency of method for OA detection. The results shows acceptable recovery rate and low RSD, demonstrating the applicability of this method for in suit OA assessment in the quality and toxicity monitoring of seafood.

It is an undeniable fact that the effect of marine toxins becomes increasingly serious to people. Paralytic shellfish poisons (PSP) are well-known sodium channel-blocking marine toxins, which block the conduction of nerve impulses and lead to a series of neurological disorders symptoms. Similarly, as inhibitors of serine/threonine protein phosphatases type 1 (PP1) and 2A (PP2A), diarrheic shellfish poisons (DSP) can destroy cytoskeleton and affect intracellular signal transduction, which causes some symptoms of gastrointestinal food allergy. Since the conventional methods to detect marine toxins had some disadvantages (e.g., complicated pretreatment, time-consuming detection, huge instruments and off-site test), bionic electronic eye (Bionic e-Eye) was designed and studied for the in-field fast measurement and real-time on-line analysis to marine toxins. Bionic e-Eye installed the homemade software-iPlate and completed the integration of image acquisition and further data processing. With the cooperation of ELISA, Bionic e-Eye used two color models-HSV and RGB to evaluate its bioanalytical performance in saxitoxin (STX) assay and okadaic acid (OA), which were the standard representations of PSP and DSP, respectively. Moreover, the microtiter plate reader (MTPR) was compared with Bionic e-Eye in the detection of marine toxins. The results demonstrated that Bionic e-Eye using saturation had the comparable precision, dynamic range, detection limit and sensitivity to MTPR in the two assays. Therefore, Bionic e-Eye has great potential to be a field-test platform and becomes the supplement of conventional off-site detection methods in measurement domain of marine toxins. (C) 2016 Elsevier B.V. All rights reserved.

Okadaic acid (OA) and saxitoxin (STX) are common marine toxins which can accumulate in shellfish and affect human health through the food chain. A convenient and efficient method is urgently required to perform on-site detection of marine toxins to avoid frequent poisoning incidents. In this work, a smartphone-based system cooperating with competitive immunoassay strips was designed for rapid on-site detection of OA and STX. The smartphone was employed as a light detector for image acquisition and data processing. The 3D-printed portable accessory of the smartphone was utilized to fix the test strips. The homemade APP -iStrip used the pixel scan to obtain the valley value of the pixel curve as the output of the strip. The strip analytical method was easy to operate with a detection limit of 2.800 ng ml(-1) for OA and 9.808 ng ml(-1) for STX. Moreover, with the optimization of the antigen-antibody ratio on the strip, it provided quantitative analysis of OA and STX in the range of 3-20 ng ml (-1) and 10-100 ng ml(-1), respectively. Taking advantage of the smartphone and biochemical analysis, the strip analytical system will be a promising tool for on-site rapid detection of these two toxins.

Cell-based bioassays were effective method to assess the compound toxicity by cell viability, and the traditional label-based methods missed much information of cell growth due to endpoint detection, while the higher throughputs were demanded to obtain dynamic information. Cell-based biosensor methods can dynamically and continuously monitor with cell viability, however, the dynamic information was often ignored or seldom utilized in the toxin and drug assessment. Here, we reported a high-efficient and high-content cytotoxic recording method via dynamic and continuous cell-based impedance biosensor technology. The dynamic cell viability, inhibition ratio and growth rate were derived from the dynamic response curves from the cell-based impedance biosensor. The results showed that the biosensors has the dose-dependent manners to diarrhetic shellfish toxin, okadiac acid based on the analysis of the dynamic cell viability and cell growth status. Moreover, the throughputs of dynamic cytotoxicity were compared between cell-based biosensor methods and label-based endpoint methods. This cell-based impedance biosensor can provide a flexible, cost and label-efficient platform of cell viability assessment in the shellfish toxin screening fields.