CRISPR-Cas12a信号放大策略与便携式血糖仪耦合定量检测黄曲霉毒素B1

建立了CRISPR-Cas12a与便携式血糖仪耦合定量检测黄曲霉毒素B1(AFB1)的方法。体系中AFB1能够激活Cas12a的反式切割活性,激活后的Cas12a切割电极上蔗糖酶修饰的发夹探针,使得蔗糖酶游离到电极表面的溶液中。蔗糖酶催化蔗糖产生可以被血糖仪监测的响应信号,进而实现对AFB1的检测。在浓度0.001～0.1 ng/mL范围内,AFB1浓度与血糖信号呈良好的线性关系,线性方程为S=3.5+229.1c,检出限为0.3 pg/mL。该方法特异性强,适用于实际样品中AFB1的检测。     

基于简单信号读出的即时检测研究进展

即时检测(Point-of-care testing,POCT)技术作为一种简便、快速的在线分析检测手段,在疾病诊断、健康管理、环境监测和应急响应分析等领域得到了广泛应用.有效构建适于各种目标分析物的简单信号读出的产生、转移、放大和增敏策略,发展具有简单、快速、灵敏的分析检测信号,已成为POCT研究领域关注的热点.一些...     

磁性纳米标记检测技术在生物医学即时检测领域的研究进展

基于磁性纳米材料的磁标记检测技术具有灵敏度高、线性范围广、信号检测便捷等优点。由于生物样品自身磁背景信号极低,相比于光学标记检测技术,磁标记检测技术在蛋白质、核酸、细胞、病原体及生物组织检测中均表现出更高的灵敏度,在生物医学即时检测领域展现了良好的应用前景。该文围绕磁性纳米粒在即时检测领域的最新研究进展,重点介绍了其在蛋白质、核酸以及几类病原体检测方面的应用,并对基于磁性纳米粒的即时检测技术发展方向及应用前景进行了展望。     

基于粘度的现场快速检测方法及应用

即时检测技术（POCT）是一种价格低廉、响应灵敏、无需大型仪器、可快速得到检测结果的检测手段,通常由具有便携式“样品进-信号出”功能的生物传感器完成。虽然基于刺激响应聚合物溶胶-凝胶转变构建的生物传感器已有大量的研究报道,开发制备简单、检测快速、灵敏度高、选择性好的生物传感器依然具有十分重要的意义。近年来,研究者开发了基于聚合物粘度变化的新型生物传感器,可以一定程度上克服传统水凝胶传感器的缺点,具有较大的发展潜力。本文主要从检测平台以及输出信号模式两个方面,总结归纳了基于粘度的现场快速检测方法及应用。最后,对其发展趋势进行简要的讨论,以期为基于粘度的POCT技术的后续研究和发展提供参考。     

基于分子识别的农药残留快速检测研究进展

为了提高农作物的产量和质量,农药的使用量逐年增加,导致土壤、水和农作物等的污染加剧,对环境和人类健康造成了严重的威胁。因此,对于农药残留进行快速、灵敏的检测至关重要。近年来,多种用于农药残留快速检测的技术和产品被开发。该综述对多种识别方式在农药检测中的进展进行了介绍,包括以蛋白质和适配体为代表的生物识别、以纳米材料和大环化合物为代表的非生物识别以及基于农药独特的光学性质和化学性质实现的直接识别。最后对农药残留的快速检测进行了展望,以期为农药的即时监测（POCT）提供研究思路和方向。     

杂交链式反应介导的发光银纳米簇系统用于miRNA-21的信号放大检测

MicroRNAs(miRNAs)是多种疾病的生物标志物,同时也能为疾病治疗提供潜在靶点,因此,发展简单、快速、灵敏的miRNAs分析方法具有十分重要的意义。本研究结合杂交链式反应(HCR)高的信号放大能力和以DNA为模板合成的银纳米簇(DNA-AgNCs)优异的发光性能,构建了一种免标记的通用型荧光传感器,实现了miRNA-21的快速灵敏检测。将合成银纳米簇(AgNCs)的DNA模板封闭在HCR的反应物(发夹DNA)中,当存在靶标DNA时,发夹DNA的杂交链式组装反应被引发,释放出大量自由的AgNCs模板序列,进而引发近红外荧光DNA-AgNCs的合成,AgNCs的近红外荧光信号强度与引发链DNA的浓度成正相关。进一步通过在检测系统中引入一个封闭有HCR引发链序列的辅助发卡序列,建立通用型HCR-AgNCs传感分析系统,用于靶核酸分子检测。以miRNA-21为模型分析物,只在miRNA-21存在时,此辅助发卡才能被打开,并生成自由的HCR引发链,进而引发HCR反应和AgNCs的合成。本方法检测miRNA-21的线性范围为250 pmol/L～8 nmol/L,线性方程为(F-F     

基于核酸外切酶Ⅲ信号放大技术的荧光适配体传感器用于卡那霉素的检测

建立了一种基于核酸外切酶Ⅲ(Exo Ⅲ)和碳纳米颗粒(CNPs)的信号放大体系用于卡那霉素(KAN)检测的新方法。合成了水溶性的CNPs,并设计合成了不同序列的DNA,具体包括:卡那霉素适配体(Apt),羧基荧光素标记的信号DNA探针(FAM-DNA)和互补链cDNA。当体系中不存在KAN时,Apt与cDNA可以杂交形成双链DNA,体系中FAM-DNA处于单链状态,Exo Ⅲ不能水解单链DNA;此时,体系中加入CNPs,单链FAM-DNA被CNPs吸附,荧光发生淬灭;在KAN存在下,Apt与其靶标KAN特异性结合,此时FAM-DNA与cDNA杂交形成双链DNA,由于CNPs对双链DNA吸附较弱,DNA探针的荧光不发生淬灭。ExoⅢ可以特异性的从3’-端对FAM-DNA降解,释放FAM荧光团和cDNA,该体系通过"降解-杂交"循环,最终释放出大量的FAM荧光团。由于CNPs对FAM具有较低的亲和力,释放出的FAM不能吸附在CNPs表面,FAM荧光不会发生淬灭,实现荧光信号放大扩增作用。方法线性范围为50～100 nmol/L,检测限为2.5 nmol/L。该方法可用于实际样品牛奶中卡那霉素的检测。     

基于核酸切割酶与脱氧核酶的荧光循环放大系统检测铅(Ⅱ)

利用核酸切割酶(Nicking endonuclease)识别特定DNA双链并切割其中某条单链的性质,构建了基于8-17E脱氧核酶(8-17E DNAzyme)的pb2+荧光循环放大检测方法.pb2+可激活8-17E脱氧核酶水解RNA底物,产生并释放出的单链与分子信标探针( Molecular beacon,MB)杂交,导致其茎环结构被破坏,荧光信号恢复;同时形成含有核酸切割酶Nt.BbvCI识别位点的双链区域.在核酸切割酶Nt.BbvCI的作用下,分子信标探针被切割释放,游离出来的单链可与其它分子信标重新杂交,从而触发下一轮酶切,引起荧光检测信号的循环放大.本方法避免了8-17E脱氧核酶与底物链的修饰,最低可以检测出水溶液中1.0×10-10 mol/L Pb2+,并在2倍浓度的Zn2+,以及5倍浓度的其它干扰金属离子存在的情况下对pb2+显示出良好的选择性.本方法对环境水样中pb2+的标准加样回收率为96.1％～108.0％.     

Personal Glucose Meter for α-Glucosidase Inhibitor Screening Based on the Hydrolysis of Maltose

As a key enzyme regulating postprandial blood glucose, α-Glucosidase is considered to be an effective target for the treatment of diabetes mellitus. In this study, a simple, rapid, and effective method for enzyme inhibitors screening assay was established based on α-glucosidase catalyzes reactions in a personal glucose meter (PGM). α-glucosidase catalyzes the hydrolysis of maltose to produce glucose, which triggers the reduction of ferricyanide (K₃[Fe(CN)₆]) to ferrocyanide (K₄[Fe(CN)₆]) and generates the PGM detectable signals. When the α-glucosidase inhibitor (such as acarbose) is added, the yield of glucose and the readout of PGM decreased accordingly. This method can achieve the direct determination of α-glucosidase activity by the PGM as simple as the blood glucose tests. Under the optimal experimental conditions, the developed method was applied to evaluate the inhibitory activity of thirty-four small-molecule compounds and eighteen medicinal plants extracts on α-glucosidase. The results exhibit that lithospermic acid (52.5 ± 3.0%) and protocatechualdehyde (36.8 ± 2.8%) have higher inhibitory activity than that of positive control acarbose (31.5 ± 2.5%) at the same final concentration of 5.0 mM. Besides, the lemon extract has a good inhibitory effect on α-glucosidase with a percentage of inhibition of 43.3 ± 3.5%. Finally, the binding sites and modes of four active small-molecule compounds to α-glucosidase were investigated by molecular docking analysis. These results indicate that the PGM method is feasible to screening inhibitors from natural products with simple and rapid operations.     

Dose‐Dependent Response of Personal Glucose Meters to Nicotinamide Coenzymes: Applications to Point‐of‐Care Diagnostics of Many Non‐Glucose Targets in a Single Step

We report a discovery that personal glucose meters (PGMs) can give a dose‐dependent response to nicotinamide coenzymes, such as the reduced form of nicotinamide adenine dinucleotide (NADH). We have developed methods that take advantage of this discovery to perform one‐step homogeneous assays of many non‐glucose targets that are difficult to recognize by DNAzymes, aptamers, or antibodies, and without the need for conjugation and multiple steps of sample dilution, separation, or fluid manipulation. The methods are based on the target‐induced consumption or production of NADH through cascade enzymatic reactions. Simultaneous monitoring of the glucose and L ‐lactate levels in human plasma from patients with diabetes is demonstrated and the results are comparable to those from current standard test methods. Since a large number of commercially available enzymatic assay kits utilize NADH in their detection, this discovery will allow the transformation of almost all of these clinical lab tests into POC tests that use a PGM.     

运用金-银核壳结构纳米棒的表面增强拉曼效应检测超痕量葡萄糖

运用种子生长法合成金纳米棒溶胶,再经过化学还原法合成金-银核壳纳米棒溶胶作为表面增强拉曼活性基底。该基底可以检测低至10-18mol/L的罗丹明6G,其增强因子高达1011,具有优异的SERS增强效果。利用此基底的表面增强拉曼散射效应检测10-14~10-8mol/L的葡萄糖,并得到表面增强拉曼信号强度与相应葡萄糖浓度的工作曲线。实验结果表明,该基底在葡萄糖的检测及半定量分析方面具有良好的潜在应用前景。     

Gold Nanoparticle‐Based Redox Signal Enhancement for Sensitive Detection of Human Chorionic Gonadotropin Hormone

A sensitive immunosensor for the detection of pregnancy marker, human chorionic gonadotropin hormone (hCG), was developed using the direct electrical detection of Au nanoparticles. We utilized disposable screen‐printed carbon strips (SPCSs) for the development of our immunosensor, which provided cost‐effective tests with the required antigen sample volume as small as 2 μL. After the recognition reaction between the surface‐immobilized primary antibody and hCG, the captured antigen was sandwiched with a secondary antibody that was labeled with Au nanoparticles. Au nanoparticles were exposed to a preoxidation process at 1.2 V for 40 s, which was subsequently followed with a reduction scan on the same surface using differential pulse voltammetry (DPV). We could observe Au nanoparticle‐labeled antigen‐antibody complexes immobilized on the surface of SPCS using scanning electron microscopy (SEM). Additionally, the number of Au nanoparticles on the immunosensor was determined using SEM images, and showed a linear relationship with the current intensity obtained from the DPV measurements with a detection limit of 36 pg/mL hCG (612 fM, 3.6×10^?4 IU/mL). Our immunosensor system, a combination of the screen‐printing technology with Au nanoparticles provides a promising biosensor for various applications in life sciences.     

方波脉冲法制备Cu_2O@Cu/TiO_2三维阵列纳米复合材料及其作为高灵敏无酶型葡萄糖传感器的研究

采用阳极氧化法制备了TiO_2三维阵列纳米管(TiO_2 NTAs),再利用方波脉冲法在TiO_2 NTAs表面电沉积Cu薄膜,经过煅烧处理后得到Cu_2O@Cu/TiO_2 NTAs纳米复合材料。X-射线衍射仪(XRD)测试表明该材料中Cu和Cu_2O共存。扫描电镜(SEM)和透射电镜(TEM)分析显示,TiO_2管壁的外表面均匀生长Cu_2O,选区电子衍射结果表明该纳米复合材料为混晶材料。差分脉冲伏安法表明,Cu_2O@Cu/TiO_2 NTAs在+0.56 V处出现明显的葡萄糖氧化峰,表明该纳米复合材料对葡萄糖具有强的电催化作用,可用于葡萄糖的测定。计时电流法得到葡萄糖的线性范围为0.19～3.5 mmol/L,灵敏度为372.0μA·L·mmol~(-1)·cm~(-2),检出限为3μmol/L。交流阻抗测得Cu_2O/TiO_2 NTAs的传荷阻抗为526.2Ω,Cu_2O@Cu/TiO_2 NTAs为1.8Ω,二者相差292倍,说明Cu作为中间过渡层降低了电子传导的传荷阻抗。将该电极用于血清中葡萄糖含量的检测,测试结果与医院测定结果无显著差异。Cu_2O@Cu/TiO_2 NTAs可作为一种灵敏度高、响应速度快、选择性较好的无酶型葡萄糖传感器。     

Imidazolium Based Surface Active Ionic Liquids as Novel Micellar Media for Simultaneous and Sensitive Electrochemical Detection of Dopamine and Ascorbic Acid

Surface active ionic liquid (SAIL) micelle assisted, simultaneous and highly sensitive electrochemical sensing of dopamine (DA) and ascorbic acid (AA) is presented. Results presented herein establish that SAILs viz.1‐dodecyl‐3‐methyl imidazolium chloride ([DDMIM][Cl]), 1‐octyl‐3‐methyl imidazolium chloride ([OMIM][Cl]) and 1‐butyl‐3‐methyl imidazolium chloride ([BMIM][Cl]) exhibit a probe and SAIL nature/concentration specific impact on the redox behaviour of hydroquinone (H₂Q), dopamine (DA) and ascorbic acid (AA). To our observations, the electrochemical behaviour of DA and AA is affected oppositely by SAILs with the apparent effects being more appreciable in presence of [DDMIM][Cl]. In the presence of [DDMIM][Cl] micelles, the electro‐oxidation of AA was observed to occur at potentials about 350 mV less positive than required for electrooxidation of DA, an important advantage that minimises the interference of former in sensing of the later. The peak to peak potential separation of 350 mV observed in presence of [DDMIM][Cl] micelles is the largest to be reported so far. The DPV signal for DA and AA displayed a linear response in the concentration range of 6.6 to 99.9 μM and 6.6 to 131.5 μM respectively. Very low detection limits of 0.0161 μM for DA in presence of 39.8 μM AA and 0.0227 μM for AA in presence of 39.8 μM DA were estimated in micellar phase of [DDMIM][Cl].     

导电聚合物喷雾电离质谱法对微量全血代谢轮廓即时监测

血液代谢物水平可用于术后生理代谢状况的个体化精准评估,这对体液整体代谢轮廓的即时检验提出了迫切的技术需求.该文介绍了导电聚合物喷雾电离质谱(CPSI-MS)在体液代谢物多重监测中的应用价值.该技术采用由碳纳米管(CNT)和聚甲基丙烯酸甲酯(PMMA)组成的导电聚合物作为血液加载与原位电离喷头.当对该聚合物施加4.5 k...     

Synthesis and Characterization of Novel Biodegradable Di‐ and Tri‐Block Copolymers Based on Ethylene Carbonate Polymer as Hydrophobic Segment

Synthesis of novel amphiphilic biodegradable block copolymers based on ethylene carbonate is reported in this study. Polyethylene glycol monomethyl ether (MeO‐PEO) and polyethylene glycol (PEG) of varying molar masses are used as macro‐initiator for ring‐opening polymerization of ethylene carbonate in the presence of sodium stannate trihydrate as a heterogeneous transesterification catalyst. Earlier elution of block copolymer from macro‐initiator in size exclusion chromatography (SEC) indicated the successful synthesis of the block copolymers. Ratios of both types of blocks are varied systematically. Liquid chromatography at critical conditions is used for the analysis of the non‐critical individual blocks, and if there are any critical segments that are not attached to the non‐critical block. To the best of our knowledge, this is the first report on the synthesis of ethylene carbonate‐based amphiphilic block copolymers. Chromatographic critical conditions of the ethylene carbonate polymer are also reported for the first time. © 2017 Wiley Periodicals, Inc. J. Polym. Sci., Part A: Polym. Chem. 2017 , 55, 1887–1893