基于DNA长距自组装的电化学方法用于废水中痕量铽离子检测

本文基于G-四链体结构和DNA长距自组装技术,构建一种非标记型电化学交流阻抗方法用于铽离子的痕量检测。首先,根据Tb~(3+)诱导单链DNA折叠成G-四链体的性质,设计一组捕获探针(Cp)和信号探针(Tp)序列,组装到电极表面,当Tb~(3+)存在时,诱导Tp从双链结构中解离并折叠成G-四链体,导致电极表面的Cp裸露,进而与辅助探针Ap1、Ap2进行级联式杂交,在电极表面形成长距"三明治"DNA结构,通过电化学交流阻抗技术检测Tb~(3+)加入前后Ret值的变化,实现对Tb~(3+)的超高灵敏检测。在1.0×10~(-15)M~1.0×10~(-12)M范围内,?Ret与Tb~(3+)浓度的对数呈良好的线性关系,方法检测限为3.4×10~(-16)M。将其应用于实际样品中铽离子的痕量检测,结果令人满意。     

Ba2+共掺杂YPO4∶Tb3+荧光材料的水热合成与荧光性能

采用水热法合成Ba~(2+)共掺杂YPO4∶Tb~(3+)荧光材料,并通过X射线衍射(XRD)、扫描电子显微镜(SEM)和荧光分光光度计等研究了合成样品的物相组成和荧光性能,并分析了Ba~(2+)掺杂量和反应体系pH值等对合成样品的物相结构及荧光性能的影响。结果表明,反应体系pH值和Ba~(2+)掺杂量直接影响所制备样品的结构与性能。少量Ba~(2+)(≤10%,原子分数,下同)共掺杂YPO4∶1%Tb~(3+)样品均为纯相四方晶系磷钇矿结构晶体,过量Ba~(2+)掺杂导致Ba_3(PO_4)_2杂质相的出现;pH值为6的水热环境下可获得高结晶度的单一相Ba~(2+)、Tb~(3+)共掺杂YPO4样品。激发和发射光谱测试结果表明,所制备的YPO4∶1%Tb~(3+),x%Ba~(2+)样品可被225nm的紫外光有效地激发而发射出强烈的Tb~(3+)特征的黄绿色光。一定量的Ba~(2+)共掺杂可以有效地提高YPO4∶1%Tb~(3+)样品的荧光性能,但过量(高于10%)的Ba~(2+)掺杂又会导致Tb~(3+)的荧光猝灭现象出现,最佳的Ba~(2+)共掺杂量为10%。所制备的YPO4∶1%Tb~(3+),10%Ba~(2+)样品在225nm紫外光激发下位于545nm处的发射带强度是YPO4∶1%Tb~(3+)样品的1.8倍。     

ATP/CaF_2∶Ln~(3+)(Ln∶Eu,Tb,Ce/Tb)纳米粒子的制备及其荧光性能

采用两步法制备了三磷酸腺苷二钠(ATP)修饰的ATP/CaF_2∶Ln~(3+)(Ln∶Eu,Tb,Ce/Tb)纳米粒子。采用红外光谱(IR)、X-射线衍射分析(XRD)、透射电子显微镜(TEM)对所合成的纳米粒子进行结构表征,并通过荧光光谱(FS)研究了纳米粒子的荧光性能。结构研究结果表明,ATP成功地包覆在纳米粒子的表面,纳米粒子的晶相为CaF_2的立方结构,ATP/CaF_2∶Eu~(3+)、ATP/CaF_2∶Tb~(3+)、ATP/CaF_2∶Ce~(3+)/Tb~(3+)纳米粒子的平均粒径分别约为14nm、15nm、11nm。荧光性能研究表明,ATP/CaF_2∶Eu~(3+)、ATP/CaF_2∶Tb~(3+)纳米粒子基本不发射稀土离子的特征荧光,而发射出修饰剂ATP的荧光,由于Ce~(3+)对Tb~(3+)的敏化作用,ATP/CaF_2∶Ce~(3+)/Tb~(3+)纳米粒子发射出Tb~(3+)的特征荧光。     

可视化黄色荧光石油焦基碳量子点高效检测Cu~(2+)（英文）

以石油焦为碳源,采用超声辅助的化学氧化法直接制备可视化黄色荧光碳量子点(CQDs)。作为非标记的探针,该CQDs无需任何修饰即可成功用于实际水样中Cu~(2+)的检测。该荧光探针制备方法简单、经济,可快速响应(3 s),具有良好的选择性、灵敏性和可重复利用性,并且可实现"混合即检测"的快速检测目的。其线性检出范围较宽,为0.25~10μmol/L,检出限为0.029 5μmol/L。光诱导电子转移机理可很好地解释Cu~(2+)猝灭CQDs的过程,本文提出的CQDs-Cu~(2+)-EDTA"off-toon"检测机理为金属离子荧光探针的开发奠定了理论基础。石油焦基CQDs在实际水样Cu~(2+)的检测中具有快速响应性,在实际传感器应用领域具有很好的实际应用价值。     

CaWO_4∶Eu~(3+),Tb~(3+)白色荧光粉的制备及其发光性能

在不添加助剂的条件下,用微波共沉淀法法制备了铕、铽(Eu~(3+)、Tb~(3+))共掺杂的钨酸钙(CaWO_4)荧光粉。利用X射线衍射(XRD)、场发射扫描电子显微镜(SEM)、荧光光谱(PL)等表征手段,对荧光粉的物相组成、形貌和发光性能进行了分析。研究了Eu~(3+)、Tb~(3+)的掺杂比例及总掺杂量、反应温度及反应物浓度对荧光性能的影响。结果表明,Eu~(3+)、Tb~(3+)的掺杂摩尔比例、总掺杂量、温度以及反应物浓度对荧光粉的发光性能均能产生影响,其中在温度为80℃、反应物浓度为0.12mol/L且Eu~(3+)和Tb~(3+)总物质的量比金属离子总物质的量为13.1%时,得到的Eu~(3+)、Tb~(3+)共掺荧光粉在256nm激发下发射光谱色坐标为(0.270,0.236),位冷白光区。     

二元溶剂热法合成花生状CaWO_4∶Tb~(3+)及其发光性能研究

分别以水和乙醇、乙二醇的二元溶剂热法及热处理合成四方晶系CaWO_4∶Tb~(3+)球状、花生状微晶结构,利用XRD、SEM、FT-IR、DRS和PL等对其结构和形貌及性能进行表征。在不同溶剂中,反应温度为120℃,反应时间为24h的溶剂热条件下所得前驱物置于600℃煅烧后分别得到四方晶系CaWO_4∶Tb~(3+)微球、花生状微晶结构;用253nm的紫外光激发产物都发射出绿色光;花生状CaWO_4∶Tb~(3+)的荧光强度明显强于微球。     

稀土荧光玻璃

稀土荧光玻璃是由稀土元素掺杂在玻璃中,并利用稀土离子的光谱性质使基础玻璃产生可见荧光而形成的。稀土元素自由原子的基态电子组态有两种类型:[Xe]4f~n6s~2和[Xe]4f~(n-1)5d'6s~2其中[Xe]=1s~22s~22p~63s~23p~63d~(10)4s~24p~64d~(10)5s~25p~6。它们未饱和的4f电子,在紫外,可见等高能射线的激发下,从基态跃迁到激发态,然后再从激发态返回到能量较低的能态时,放出辐射能而发出荧光。在14种稀土元素中,Sm~(3+)、Eu~(3+)、Tb~(3+)、Dy~(3+)等离子,其激发态往往与配体的三     

罗丹明B多层荧光纳米微球对Hg~(2+)的检测

利用水热法合成了掺杂锌的碲化镉量子点(CdTe∶Zn QDs),并采用反相微乳法将SiO_2包覆在量子点表面形成多层SiO_2纳米微球。通过接枝方法将罗丹明B衍生物连接在硅球上,利用Hg~(2+)可以使罗丹明B衍生物的螺环结构发生开环反应并产生荧光增强这一特性,构建了荧光共振能量转移(FRET)的比率探针,并成功实现了对水溶液中Hg~(2+)的比率检测。结果表明,该探针在波长为521nm和577nm时对Hg~(2+)具有良好的灵敏度和选择性,检测限是0.5μM。     

基于铝酸盐类长余辉材料的Fe3+荧光探针

SrAl_2O_4∶Eu~(2+),Dy~(3+)是常见的长余辉发光材料,具有良好的发光和余辉性能,能够作为荧光共振能量转移(FRET)体系中能量供体,使探针具备在无光源激发条件下的检测能力.但其易发生水解反应而失去光学性能,这限制了其应用.对SrAl_2O_4∶Eu~(2+),Dy~(3+)进行表面修饰改性,并以其为基体制备可利用余辉进行检测的荧光探针具有重要的研究意义.本文利用正硅酸乙酯(TEOS)在酸性条件下的水解缩合在SrAl_2O_4∶Eu~(2+),Dy~(3+)表面包覆SiO_2以提高其耐水性,并通过硅烷偶联剂将其与罗丹明B连接构成一种新型荧光探针.采用扫描电镜、荧光光谱等检测手段对所制备探针进行表征,并分别研究了探针在有、无光源激发条件下的检测性能.结果表明,制备的探针对Fe~(3+)有较特殊的响应,可用于检测溶液中的Fe~(3+).该探针在光源激发下可定量检测溶液中100～500μmol/L浓度范围内的Fe~(3+),在无光源激发条件下可利用自身余辉检测溶液中500μmol/L以上浓度的Fe~(3+).     

基于荧光标记的液相芯片检测方法研究

讨论了基于荧光标记的液相芯片及其二维并行检测方法. 设计了待测试液的特殊流场. 利用脉冲激光激发微球探针上的荧光,配合高灵敏度的 CCD 检测荧光信息,计算机对各个 CCD 记录的荧光信息进行后期处理,获得液相芯片中微球探针上的全部信息. 对检测技术方案中探针流态和荧光信号的强度进行了分析,建立了荧光信号强度与检测系统中各参量间的表达式. 每个探测单元都对荧光信号的收集进行了验证实验. 这种检测方法具有快速、准确、灵敏度高的优点.     

Sensitive and homogeneous protein detection based on target-triggered aptamer hairpin switch and nicking enzyme assisted fluorescence signal amplification

Specific and sensitive detection of proteins in biotechnological applications and medical diagnostics is one of the most important goals for the scientific community. In this study, a new protein assay is developed on the basis of hairpin probe and nicking enzyme assisted signal amplification strategy. The metastable state hairpin probe with short loop and long stem is designed to contain a protein aptamer for target recognition. A short Black Hole Quencher (BHQ)-quenching fluorescence DNA probe (BQF probe) carrying the recognition sequence and cleavage site for the nicking enzyme is employed for fluorescence detection. Introduction of target protein into the assay leads to the formation change of hairpin probe from hairpin shape to open form, thus faciliating the hybridization between the hairpin probe and BQF probe. The fluorescence signal is amplified through continuous enzyme cleavage. Thrombin is used as model analyte in the current proof-of-concept experiments. This method can detect thrombin specifically with a detection limit as low as 100 pM. Additionally, the proposed protein detection strategy can achieve separation-free measurement, thus eliminating the washing steps. Moreover, it is potentially universal because hairpin probe can be easily designed for other proteins by changing the corresponding aptamer sequence.     

Polymeric sequence probe for single DNA detection

A polynucleotide probe, polymeric sequence probe (PSP), was developed for single molecular detections. PSP is a single-stranded DNA molecule with ~2000 tandem repeat target-binding sequences and label-binding sequences. A single PSP can bind to multiple fluorescent complementary oligos to generate a strong fluorescence signal. Single target molecules bound to PSPs can be clearly visualized by a conventional fluorescence microscope. An ultrasensitive PSP-based assay for Mycobacterium tuberculosis was demonstrated.     

新型荧光定量模型用于湘江水中铅离子的检测

富G序列AGRO100核酸探针与荧光染料硫磺素T（ThT）和N-甲基卟啉二丙酸IX（NMM）结合后,随着铅离子的加入ThT和NMM的荧光强度会发生相对变化,从而建立了用于铅离子定量分析的、免标记的广义比率型荧光传感方法。将该方法与新型荧光定量模型（QFM）相结合,分析4个不同地段湘江水中铅离子的浓度。研究结果表明：广义比率型荧光探针结合QFM,能够准确定量分析4个不同地段湘江水中铅离子的浓度,该分析结果与电感耦合等离子体质谱仪（ICP-MS）得到的结果基本一致,加标回收率均在91%~108%的范围内。     

Monitoring molecular beacon/DNA interactions using atomic force microscopy

The molecular beacon (MB) is a new fluorescence probe containing a single-stranded oligonucleotide with a probe sequence embedded in complementary sequences that form a hairpin stem. Due to the inherent fluorescent signal transduction mechanism, an MB functions as a sensitive probe with a high signal-to-background ratio for real-time monitoring and provides a variety of exciting opportunities in DNA, RNA, and protein studies. To better understand the properties of MBs, the specific interactions between MB and target DNA (complementary and one-base mismatch) have been directly investigated by atomic force microscopy. The interaction force between a linear DNA probe and the target DNA was also detected and compared to that between MB and target DNA. The results demonstrate the high specificity of the MB/target DNA compared to the linear DNA/target DNA interaction.     

Probes for detection of specific DNA sequences at the single-molecule level

A method has been developed for highly sensitive detection of specific DNA sequences in a homogeneous assay using labeled oligonucleotide molecules in combination with single-molecule photon burst counting and identification. The fluorescently labeled oligonucleotides are called smart probes because they report the presence of complementary target sequences by a strong increase in fluorescence intensity. The smart probes consist of a fluorescent dye attached at the terminus of a hairpin oligonucleotide. The presented technique takes advantage of the fact that the used oxazine dye JA242 is efficiently quenched by complementary guanosine residues. Upon specific hybridization to the target DNA, the smart probe undergoes a conformational change that forces the fluorescent dye and the guanosine residues apart, thereby increasing the fluorescence intensity about six fold in ensemble measurements. To increase the detection sensitivity below the nanomolar range, a confocal fluorescence microscope was used to observe the fluorescence bursts from individual smart probes in the presence and absence of target DNA as they passed through the focused laser beam. Smart probes were excited by a pulsed diode laser emitting at 635 nm with a repetition rate of 64 MHz. Each fluorescence burst was identified by three independent parameters: (a) the burst size, (b) the burst duration, and (c) the fluorescence lifetime. Through the use of this multiparameter analysis, higher discrimination accuracies between smart probes and hybridized probe-target duplexes were achieved. The presented multiparameter detection technique permits the identification of picomolar target DNA concentrations in a homogeneous assay, i.e., the detection of specific DNA sequences in a 200-fold excess of labeled probe molecules.     

Hairpin fluorescence DNA probe for real-time monitoring of DNA methylation

DNA methylation catalyzed by methylase plays an important role in many biological events. However, traditional methods of methylase activity analysis by gel electrophoresis were laborious and discontinuous. In this paper, we report a new strategy to study methylase activity using fluorescent probes coupled with enzyme-linkage reactions. A hairpin DNA probe is prepared with a fluorophore and a quencher linked at the 5'- and 3'-terminus of the probe. A disturbance of the stem sequence by DNA methylation would cause the separation of the fluorophore and the quencher, resulting in the restoration of the fluorescence. We used DNA adenine methylation (Dam) methyltransferase (MTase) and Dpn I endonuclease, both having a 5'-G-A-T-C-3' recognition sequence. Dam MTase catalyzed the methylation of the sequence of 5'-GATC-3', and Dpn I cut the sequence of 5'-G-Am-T-C-3'. The fluorescence of the hairpin probe was restored when it was cleaved by Dpn I endonuclease during the course of methylation. Unlike traditional methods, this assay was done in real time and could be used to monitor the dynamic process of methylation. Our method is easy, simple, and nonradioactive, yet as efficient as gel electrophoresis in detecting the activity of methylase. It also had the potential to screen suitable inhibitor drugs for Dam methylase.     

催化发夹组装用于碱基切除修复酶活性检测研究

碱基切除修复酶在 DNA 损伤修复过程中具有重要作用,且其检测 与癌症等疾病的诊断相关。传统的碱基切除修复酶检测方法操作复杂、灵敏 度低,且仅对酶的浓度进行定量分析而非酶的活性。为此,建立了基于催化 发夹组装介导信号放大用于核酸内切酶 IV（Endo IV）活性的检测方法。该 方法利用 Endo IV 的活性作用于底物探针,将引发序列释放而引发催化发夹 自组装信号放大,以实现 Endo IV 的活性检测分析。通过荧光检测实验可知, 该方法检测下限为 3.7×10 -7 U/mL,可选择性地对 Endo IV 的活性进行检测, 是一种设计简单、操作简便、灵敏度高的碱基切除修复酶活性检测方法。     

Fluorescence turn-on detection of DNA and label-free fluorescence nuclease assay based on the aggregation-induced emission of silole

By making use of the aggregation-induced emission feature of silole, compound 1 with an ammonium group is designed and synthesized with a view to developing a new optical probe for fluorescence turn-on detection of DNA and label-free fluorescence nuclease assay. The fluorescence of 1 increases largely upon mixing with DNA, in particular for long DNA, indicating that 1 can be used for fluorescence turn-on detection of DNA. More interestingly, 1 can be employed to follow the DNA cleavage process by nuclease. Therefore, a label-free fluorescence nuclease assay method is successfully established with 1. Furthermore, this label-free fluorescence assay can also be used for inhibitor screening of nucleases. Given its simplicity, easy operation, sensitivity and cost-effectiveness, this method can be extended to other nuclease assays and high-throughput screening of nuclease inhibitors.     

Double-strand DNA-templated formation of copper nanoparticles as fluorescent probe for label-free aptamer sensor

Double-strand DNA (dsDNA) can act as an efficient template for the formation of copper nanoparticles (Cu NPs) at low concentration of CuSO(4), and the formed Cu NPs have excellent fluorescence, whereas a single-strand DNA (ssDNA) template does not support Cu NPs' formation. This property of dsDNA-Cu NPs makes it suitable for DNA sensing. However, exploration of dsDNA-Cu NPs applied in biological analysis is still at an early stage. In this regard, we report herein for the first time a sensitive, cost-effective, and simple aptamer sensor (aptasensor) using dsDNA-Cu NPs as fluorescent probe. The design consists of a dsDNA with reporter DNA (here, aptamer) as template for the formation of Cu NPs, and the formed dsDNA-Cu NPs show high fluorescence. Using adenosine triphosphate (ATP) as a model analyte, the introduction of ATP triggers the structure switching of reporter DNA to form aptamer-ATP complex, causing the destruction of the double helix and thus no formation of the Cu NPs, resulting in low fluorescence. The preferable linear range (0.05-500 μM), sensitivity (LOD 28 nM), and simplicity for the detection of ATP indicate that dsDNA-Cu NPs may have great prospects in the field of biological analysis. We also use this novel fluorescent probe to determine ATP in 1% human serum and human adenocarcinoma HeLa cells. The dsDNA-Cu NPs probes provide recovery of 104-108% in 1% human serum and a prominent fluorescent signal is obtained in cellular ATP assay, revealing the practicality of using dsDNA-Cu NPs for the determination of ATP in real samples. Besides, this design is simply based on nucleic acid hybridization, so it can be generally applied to other aptamers for label-free detection of a broad range of analytes. Successful detection of cocaine with detection limit of 0.1 μM demonstrates its potential to be a general method.     

Butterfly-shaped conjugated oligoelectrolyte/graphene oxide integrated assay for light-up visual detection of heparin

A water-soluble pyrene-based butterfly shaped conjugated oligoelectrolyte (TFP) is synthesized and integrated with graphene oxide (GO) to form a label-free assay for heparin detection. Efficient fluorescence quenching occurs between TFP and GO because of strong electrostatic and π-π interactions, leading to nearly dark emission in the absence of analytes. Addition of heparin into TFP solution significantly minimizes the fluorescence quenching of GO toward TFP, which is less effective for the heparin analogues, such as hyaluronic acid and chondroitin 4-sulfate. As a consequence, the solution emits strong yellow fluorescence only in the presence of heparin, which allows for light-up visual discrimination of heparin from its analogues. Moreover, the linear light-up response of the TFP/GO integrated assay enables heparin quantification in the range of 0-1.76 U/mL with a limit of detection of 0.046 U/mL, which is practical for heparin monitoring during postoperative and long-term care. This study thus demonstrates a new synthetic strategy to develop GO-based chemical and biological sensing without the employment of dye-labeled biomolecules.