一种基于DNA G-四链体结构的汞离子快速检测方法

随着工业的发展汞污染日渐严重,建立一种快速、准确的Hg²⁺检测手段变得更加重要。某些富鸟嘌呤(G)的核酸序列能够通过氢键和自身链或者其他序列链上的鸟嘌呤(G)连接折叠成为四链结构,称为G-四链体。Hg²⁺可以和G-四链体中的胸腺嘧啶T相互作用形成T-Hg²⁺-T结构,G-四链体也会形成DNA双链结构,这种变化可以影响菁染料的聚集状态,进而引起溶液紫外-可见吸收光谱的变化。基于DNA G-四链体转化菁染料聚集状态开发了一种Hg²⁺检测手段。     

链置换反应触发G-四链体DNA酶双向自组装用于高灵敏均相比色生物传感

结合高特异性核酸适配体识别及链置换作用触发杂交链反应,成功实现了大量G-四链体DNA酶活性结构的双向自组装,进一步利用该DNA酶优良的类过氧化物酶性质发展了一种凝血酶(Tb)均相比色生物传感新方法。基于链置换辅助的目标物循环作用以及杂交链反应引起的DNA酶双向自组装信号放大,该方法可在3个数量级线性范围内实现对Tb的高灵敏检测,检测限为0.82 pmol·L~(-1)。     

硝基取代钌配合物-适配体荧光探针检测汞离子

生物体内汞离子过量会对神经系统和正常的代谢活动造成极大的伤害.因此有必要发展新的方法用于汞离子的高灵敏、高选择性检测.本研究采用钌配合物-适配体探针体系,利用钌配合物可选择性识别G-四链体DNA以及核酸适配体中T碱基与汞离子特异性结合的特性,建立了一种荧光检测汞离子的方法.当无汞离子时,核酸适配体形成G-四链体结构;有...     

基于末端脱氧核苷酸转移酶协同G-四链体核酶的恩诺沙星检测新方法

恩诺沙星因抗菌活性强、抗菌谱广而广泛应用于动植物疾病治疗，但过度使用甚至滥用会导致其在动植物食品中的残留量超标，因此发展高效的恩诺沙星检测方法至关重要。本工作基于末端脱氧核苷酸转移酶（TdT）协同G-四链体核酶设计信号放大策略，建立了一种新型恩诺沙星（ENR）电化学检测方法。目标物恩诺沙星与特异性核酸适体的结合触发TdT在电极表面的扩增反应，产生G-四链体核酶纳米线结构，进而发挥辣根过氧化物酶活性催化信号放大，实现恩诺沙星的高灵敏和高特异性检测。本方法对恩诺沙星的线性检测范围为0.5 ~ 50 ng/mL，检测限低至0.043 ng/mL。此外，该无标记电化学生物传感器简单快速，成本低，并成功应用于对实际食品样本的分析检测，显示出较好的应用潜能。     

G-四联体的小分子配体的研究进展

位于染色体末端(如端粒)以及内染色体区域(致癌启动子区)的G-四联体结构因涉及癌变基因(c-myc、c-kit、K-ras等)的基因表达以及末端区域的结构稳定性,并且能够通过端粒酶来抑制端粒的延长(不会缩短的端粒可以使癌细胞无限增殖),逐渐成为抗癌药物的靶点。作为G-四联体的配体,自然界中分布广泛的药物因具有多态性、结构复杂性以及潜在的活性成为首选。本文综述自然药物作为G-四联体的配体以及其优越的抗肿瘤活性。     

研究G-四链体与其配体相互作用的技术方法概述

近年来以G-四链体DNA为药物靶点的研究引起了人们的广泛关注,G-四链体与其配体相互作用的研究方法也多种多样.本文对研究G-四链体形成的结构,配体与G-四链体结合能力,配体与G-四链体结合模式等三个方面的技术方法进行了综述.     

基于3D打印技术构建石墨烯电化学生物传感器及其应用研究

超高灵敏度和特异性的电化学生物传感器在环境风险物质监测以及生物医学检测领域具有重要意义,而构建生物亲和性高、制备工艺简单、成本低廉的检查电极是电化学生物传感器走向应用的关键。本文采用3D打印技术制备出重复性良好的三维石墨烯复合电极,然后通过电化学氧化的方法调控表面石墨烯的形貌和氧化基团。所制备的电化学生物传感器在环境污染物微囊藻毒素(MC-LR)的检测中展现出超高的灵敏度,其线性检测区在4×10^-6~1 μg/L,检测限为1.5×10^-7 μg/L。同时,通过改变适配体检测探针后,该电化学生物传感器对于多巴胺、重金属Hg²⁺、四环素等均具有极高的检测灵敏度。本研究为电化学适配体生物传感器走向应用化提供了一种新的思路,为开发超高灵敏度环境监测和生物医学检测传感器提供一定的基础数据。     

利用功能性DNA结构对重金属铅离子进行快速检测

重金属铅离子是具有较大毒性并且属于严重破坏环境的污染物,铅离子的快速检测受到了人们的广泛关注。采用功能性DNA和菁染料ETC的特异性识别构建铅离子探针,达到快速响应检测铅离子含量的目的。菁染料ETC能够特异性识别平行构型G-四链体,在吸收光谱上表现出单体特征吸收峰,铅离子能够将平行构型G-四链体逐渐诱导形成反平行G-四链体,从而引起ETC的聚集体变化,根据特征吸收峰值的变化来检测铅离子含量。     

适配体传感器在磺胺类抗生素检测中的应用研究进展

适配体生物传感器是检测环境中抗生素含量的有效手段。介绍了磺胺类抗生素检测中常用的3种适配体生物传感器的工作原理、特点、研究现状,包括适配体电化学传感器、适配体荧光传感器和适配体比色传感器。     

异喹啉类生物碱和G-四链体结合的分子动力学研究

G-四链体是核酸的一种非经典二级结构,主要出现于富含鸟嘌呤（G）碱基的DNA或RNA序列。生物体内的G-四链体主要形成于端粒区域和某些原癌基因的启动子区域,是生物医学研究的重要对象。以G-四链体作为靶点的抗癌策略虽被多次提出,但目前还未有成功进入临床试验的案例。因此,有关原癌基因启动子区域的G-四链体与配体结合的研究,可以对靶向抗癌药物的设计提供指导性建议。使用分子动力学模拟的方法,研究了不同的异喹啉类生物碱与G-四链体的结合机理。通过考察四种异喹啉配体与G-四链体结合的过程,得到了异喹啉配体与G-四链体稳定结合的构象以及结合的主导因素。此工作从原子分子层面深化了对异喹啉类生物碱和G-四链体结合机理的微观认识,对抗癌药物设计具有指导意义。     

Gold nanoparticle-based electrochemical biosensors

The unique properties of gold nanoparticles to provide a suitable microenvironment for biomolecules immobilization retaining their biological activity, and to facilitate electron transfer between the immobilized proteins and electrode surfaces, have led to an intensive use of this nanomaterial for the construction of electrochemical biosensors with enhanced analytical performance with respect to other biosensor designs. Recent advances in this field are reviewed in this article. The advantageous operational characteristics of the biosensing devices designed making use of gold nanoparticles are highlighted with respect to non-nanostructured biosensors and some illustrative examples are commented. Electrochemical enzyme biosensors including those using hybrid materials with carbon nanotubes and polymers, sol-gel matrices, and layer-by-layer architectures are considered. Moreover, electrochemical immunosensors in which gold nanoparticles play a crucial role in the electrode transduction enhancement of the affinity reaction as well as in the efficiency of immunoreagents immobilization in a stable mode are reviewed. Similarly, recent advances in the development of DNA biosensors using gold nanoparticles to improve DNA immobilization on electrode surfaces and as suitable labels to improve detection of hybridization events are considered. Finally, other biosensors designed with gold nanoparticles oriented to electrically contact redox enzymes to electrodes by a reconstitution process and to the study of direct electron transfer between redox proteins and electrode surfaces have also been treated.     

Single-walled carbon nanotubes as optical materials for biosensing

In this review, we summarize recent progress in the development of single-walled carbon nanotubes (SWNTs) as optical materials for biosensing applications. First, as optical labels, we discuss the use of SWNTs in Raman-based protein detection. Strong and simple resonance Raman spectroscopy of SWNTs opens up a method of protein microarray with detection sensitivity down to femtomolar range. Also, tunable isotopic SWNT-Raman signature enables the simultaneous detection of multiple analytes in complex fluids. Second, the photoluminescence properties of SWNTs are also explored. We examine fluorescence biosensors that integrate the quenching property of SWNTs and the recognition property of functional nucleic acids. Particularly, SWNTs are established as an efficient signal transduction substrate in different biosensing systems, including the detection of specific proteins and DNA sequences, regulation of singlet oxygen generation and label-free fluorescence assays, and all have exhibited very high selectivity and sensitivity.     

利用水溶性共轭高分子检测DNA的研究进展

文章介绍了利用水溶性共轭高分子的光信号放大的特点,以荧光为检测手段,对核酸进行特异性识别的生物传感器。主要介绍了DNA的两种检测方法,并研究了影响荧光共振能量传递效率的一些因素。     

Genetically Encoded Biosensors to Monitor Intracellular Reactive Oxygen and Nitrogen Species and Glutathione Redox Potential in Skeletal Muscle Cells

Reactive oxygen and nitrogen species (RONS) play an important role in the pathophysiology of skeletal muscle and are involved in the regulation of intracellular signaling pathways, which drive metabolism, regeneration, and adaptation in skeletal muscle. However, the molecular mechanisms underlying these processes are unknown or partially uncovered. We implemented a combination of methodological approaches that are funded for the use of genetically encoded biosensors associated with quantitative fluorescence microscopy imaging to study redox biology in skeletal muscle. Therefore, it was possible to detect and monitor RONS and glutathione redox potential with high specificity and spatio-temporal resolution in two models, isolated skeletal muscle fibers and C2C12 myoblasts/myotubes. Biosensors HyPer3 and roGFP2-Orp1 were examined for the detection of cytosolic hydrogen peroxide; HyPer-mito and HyPer-nuc for the detection of mitochondrial and nuclear hydrogen peroxide; Mito-Grx1-roGFP2 and cyto-Grx1-roGFP2 were used for registration of the glutathione redox potential in mitochondria and cytosol. G-geNOp was proven to detect cytosolic nitric oxide. The fluorescence emitted by the biosensors is affected by pH, and this might have masked the results; therefore, environmental CO₂ must be controlled to avoid pH fluctuations. In conclusion, genetically encoded biosensors and quantitative fluorescence microscopy provide a robust methodology to investigate the pathophysiological processes associated with the redox biology of skeletal muscle.     

Strategies for the Development of Metallic-Nanoparticle-Based Label-Free Biosensors and Their Biomedical Applications

Label-free biosensors offer accurate sensing capabilities due to the reliable quantification of biological and biochemical processes. These devices function by establishing a dynamic interaction of analyte and receptor molecules and convert this interaction into a measurable signal through a transducer. In recent decades, label-free biosensors have attracted attention in biomedical applications due to the ease of linking nanomaterials with bioreceptor molecules. In this review, recent advances in sensitivity, specificity, and sensing mechanism related to label-free biosensors of metallic nanoparticles of gold, silver, aluminium, copper, and zinc oxide are presented. Selected sensing methods based on fluorescence, surface plasmon resonance, surface-enhanced Raman scattering, metal-enhanced fluorescence, and electrochemical sensors are discussed. New measurement techniques and rapid progress of label-free biosensors are going to play a vital role in the real-time detection of biomarkers in clinical samples, such as blood plasma, serum, and urine, as well as in targeted drug delivery. Future trends of these label-free biosensing mechanisms and their development are also discussed.     

纳米CdS∶Cu双酶膜葡萄糖生物传感器

将纳米CdS∶Cu颗粒加入到葡萄糖酶(GOD)和辣根过氧化物酶(HRP)双酶膜中,与导电聚合物聚邻苯二胺(PoPD)经电化学聚合反应而固定此两酶,制备了电流型纳米CdS∶Cu颗粒双酶膜葡萄糖生物传感器,分析了CdS∶Cu纳米颗粒对传感器电流响应的影响,进行了传感器的性能测定。实验表明,引入CdS∶Cu纳米粒子和PoPD后可显著改进传感器响应性能,线性范围为0.55~9.2 mmol/L,检测下限为0.55 mmol/L,响应时间为20 s。稳定工作215天,传感器活性指标无显著变化,且抗干扰性强。     

Development of solution-gated graphene transistor model for biosensors

The distinctive properties of graphene, characterized by its high carrier mobility and biocompatibility, have stimulated extreme scientific interest as a promising nanomaterial for future nanoelectronic applications. In particular, graphene-based transistors have been developed rapidly and are considered as an option for DNA sensing applications. Recent findings in the field of DNA biosensors have led to a renewed interest in the identification of genetic risk factors associated with complex human diseases for diagnosis of cancers or hereditary diseases. In this paper, an analytical model of graphene-based solution gated field effect transistors (SGFET) is proposed to constitute an important step towards development of DNA biosensors with high sensitivity and selectivity. Inspired by this fact, a novel strategy for a DNA sensor model with capability of single-nucleotide polymorphism detection is proposed and extensively explained. First of all, graphene-based DNA sensor model is optimized using particle swarm optimization algorithm. Based on the sensing mechanism of DNA sensors, detective parameters (I_ds and V_gmin) are suggested to facilitate the decision making process. Finally, the behaviour of graphene-based SGFET is predicted in the presence of single-nucleotide polymorphism with an accuracy of more than 98% which guarantees the reliability of the optimized model for any application of the graphene-based DNA sensor. It is expected to achieve the rapid, quick and economical detection of DNA hybridization which could speed up the realization of the next generation of the homecare sensor system.     

Redox polymer-based reagentless horseradish peroxidase biosensors: Influence of the molecular structure of the polymer

Reagentless amperometric biosensors were prepared using a variety of nitrogen donor groups containing co-polymers. The polymers were coordinated with Os-bis-N,N-(2,2′-bipyridil)-dichloride via a ligand exchange reaction thus assuring an efficient electron-transfer pathway between the polymer-entrapped horseradish peroxidase and the electrode surface by means of a sequence of electron-hopping steps. The impact of structural features of the polymer such as spacer length between the Os-complex and the polymer backbone or the ratio of 4-vinylpyridine and butylmethacrylate in a co-polymer on the activity of the horseradish peroxidase biosensors was investigated.     

"DNA traffic lights": concept of wavelength-shifting DNA probes and application in an aptasensor

Add it and see it: The concept of "DNA traffic lights" for wavelength-shifting DNA probes has a great potential in the application of biosensors, for example, in DNA aptamers. A visual color change in the DNA aptasensor fluorescence from green to red occurs after specific target binding.     

Comparison of Two DNA Aptamers for Dopamine Using Homogeneous Binding Assays

Dopamine is an essential neurotransmitter and its detection is important for bioanalytical chemistry. Two very different DNA aptamers have been reported for dopamine, one derived from an RNA aptamer (named Apt1) and other obtained via direct aptamer selection (named Apt2). In this study, we used four homogeneous binding assays to compare these two DNA dopamine aptamers. Thiazole orange (TO) fluorescence assay indicated that the Apt2 specifically bound with dopamine with a K_d of 2.37 μM, which was consistent with that from the isothermal titration calorimetry (ITC) assay. However, Apt1 had much less TO fluorescence change and also no signal from ITC. By labeling the two ends of the two aptamers by a fluorophore and a quencher, the aptamer beacons showed binding of dopamine only for Apt2. Finally, the label-free AuNP-based colorimetric assay showed no difference between these two aptamer sequences, and even non-binding random DNA showed the same response, indicating that AuNPs were not a good probe for detecting dopamine. According to the data, Apt1 does not appear to be able to bind dopamine specifically, while Apt2 showed specific binding and could be used for developing related biosensors.