基于钌配合物作荧光探针测定半胱氨酸的新型DNA荧光传感器

半胱氨酸是人体内不可缺少的一种含巯基氨基酸,建立简单、灵敏测定半胱氨酸的传感器具有重要意义。本文以钌配合物Ru(phen)_2(dppx)~(2+)(phen=1,10-phenanthroline,dppx=7,8-dimethyldipyridophenazine)作为荧光探针,以银离子稳定的含错配胞嘧啶碱基的自互补DNA序列作为传感元件,利用半胱氨酸与银离子相互作用,诱导银离子稳定的双链DNA解链为单链DNA,被双链DNA保护的Ru(phen)_2(dppx)~(2+)荧光强度明显减弱,据此建立了一种测定半胱氨酸的新型DNA荧光传感器。溶液在610nm处的荧光强度与半胱氨酸的浓度在0.0～160.0 nmol/L范围内呈良好的线性关系,线性方程为:I_f=-0.122 c+57.09,相关系数(r)为0.9928,检出限(3δ/S)为13.8 nmol/L。     

以亚甲基蓝为杂交指示剂的DNA电化学传感器

构建了一个以亚甲基蓝(MB)为杂交指示剂的电化学DNA传感体系:通过自组装的方式将巯基改性的单链DNA5′端(HS-ssDNA)连接到金电极表面形成Au-ssDNA,当检测体系中加入和Au-ssDNA互补的目标单链DNA(cssDNA)时,会形成一个双链DNA系统(Au-dsDNA),加入MB并通过循环伏安法检测了杂交过程中的信号变化,验证了Au-ssDNA及Au-dsDNA的形成。在4.0×10-6～1.0×10-5mol/L内,检测灵敏度随MB浓度的增加而升高,当浓度达到2.0×10-5mol/L时接近最大值。示差脉冲伏安法检测结果表明,该检测体系对目标DNA的选择性识别能力高,可区分具有单碱基错配的目标DNA序列。检测体系对目标DNA的检测灵敏度随着目标DNA浓度的增加而增加,在5.0×10-10～1.8×10-9mol/L内呈线性关系,计算所得对目标DNA的检测限为5.0×10-10mol/L。使用寿命检测表明,经过5次变性/杂交循环后,检测信号降低并接近于检测限。     

基于荧光铜纳米簇用于钴离子的超灵敏免标记检测

为了检测浓度超标的重金属离子,探讨了一种新的荧光探针传感器检测法。利用柠檬酸钠作为还原剂合成铜纳米晶,然后加入半胱氨酸作为刻蚀剂和保护剂,与铜纳米晶配体交换反应合成荧光铜纳米簇,以铜纳米簇作为荧光探针建立了一种检测水样中重金属钴离子的方法。该方法是基于钴离子存在时,破坏铜纳米簇,导致荧光迅速降低。在较宽的钴离子浓度范围(1~100μmol/L)内,铜纳米簇荧光淬灭程度与浓度呈线性关系(R=0.970 0),其检出限较低(0.567 1μmol/L)。铜纳米簇传感器特异性选择钴离子,不受其他金属离子干扰,检测的选择性较好。基于此,完成对钴离子的超灵敏免标记检测,且EDTA可作为钴离子的螯合剂,实现了铜纳米簇传感器的循环反复使用。     

应用光敏生物素标记探针检测杂交瘤细胞中小鼠白血病病毒

采用BamHⅠ酶切和琼脂糖凝胶电泳技术，从重组质粒pSF11中回收5kb小鼠白血病病毒（MuLv）DNA片段，用光敏生物素标记制备DNA探针，采用斑点杂交法检测杂交癌细胞，McAb纯品以及SP2／0细胞中的MuLv。结果表明，此探针特异性好，灵敏度可达25pg。在检测的13株杂交瘤细胞和2株SP2／0细胞中分别有5株和1株为MuLv阳性，在McAb纯品中未查出MuLv。     

纳米粒子三明治型传感器的组装及其在BRCA-1检测中的应用

采用纳米磁、DNA、纳米金组装三明治型生物传感器。其中,纳米磁粒子(PMPS)用于捕获并分离目标DNA,而被基因探针序列修饰过的纳米金(Au-NPs)则扮演识别序列及产生信号的角色,从而使目标DNA序列量转化为光信号,继而通过紫外分光光度计进行定量分析。这种基于纳米金修饰的DNA探针的新型基因诊断技术,可以用来定量检测分析目标DNA。结果显示,这是一种非常简单经济且实用的检测单个突变基因序列的新方法。     

双信号金纳米簇对三硝基甲苯的荧光分析

三硝基甲苯作为硝基芳香化合物的代表物,应用相当广泛。其对人体健康、生活环境均具有较大的危害,因此发展简单、便捷检测三硝基甲苯的分析方法非常有必要。利用环境友好的牛血清白蛋白和2-氨基嘌呤合成了双信号金纳米簇荧光探针,基于三硝基甲苯对金纳米簇荧光的淬灭作用,建立了一种检测三硝基甲苯的荧光分析新方法,检出限可达0.14μmol/L,该方法具有良好的灵敏度和选择性。     

反应激活型酶荧光探针的研究进展

酶在维持生物体内稳态与生命活动的正常运行方面发挥着举足轻重的作用。某些特定酶含量及活性的异常与人类重大疾病的发生与发展密切相关。因此,生物体内特定酶的实时原位检测及可视化成像具有重要的意义。化学荧光探针具有选择性好、灵敏度高及高时空分辨率可视化成像等优点,近年来研究者设计合成了大量的可用于生物体系内酶识别与可视化成像的荧光探针。目前识别酶的荧光探针主要有两类:(1)基于酶对荧光探针分子中酶抑制剂基团的识别引起探针荧光信号的变化;(2)基于酶对荧光探针特异性催化反应来实现识别前后荧光信号的激活,称为反应激活型酶荧光探针。对反应激活型酶荧光探针的设计策略及4种重大疾病相关的生物标志酶(单胺氧化酶、β-半乳糖苷酶、硝基还原酶、γ-谷氨酰转肽酶)的识别可视化荧光探针研究进展进行了综述,对未来酶识别荧光探针的研究方向进行了展望。     

反应激活型酶荧光探针的研究进展

酶在维持生物体内稳态与生命活动的正常运行方面发挥着举足轻重的作用。某些特定酶含量及活性的异常与人类重大疾病的发生与发展密切相关。因此,生物体内特定酶的实时原位检测及可视化成像具有重要的意义。化学荧光探针具有选择性好、灵敏度高及高时空分辨率可视化成像等优点,近年来研究者设计合成了大量的可用于生物体系内酶识别与可视化成像的荧光探针。目前识别酶的荧光探针主要有两类：（1）基于酶对荧光探针分子中酶抑制剂基团的识别引起探针荧光信号的变化;（2）基于酶对荧光探针特异性催化反应来实现识别前后荧光信号的激活,称为反应激活型酶荧光探针。对反应激活型酶荧光探针的设计策略及4种重大疾病相关的生物标志酶（单胺氧化酶、β-半乳糖苷酶、硝基还原酶、γ-谷氨酰转肽酶）的识别可视化荧光探针研究进展进行了综述,对未来酶识别荧光探针的研究方向进行了展望。     

感染性疾病RNA病毒性病原体生物信息学检测方法的建立

目的以艾滋病病毒(HIV)为研究对象,建立感染性疾病RNA病毒性病原体的生物信息学检测方法。方法应用DNase-非序列依赖的单引物扩增技术,将艾滋病患者血清过滤后,经DNase处理去除血清中的内源DNA,提取血清病毒RNA,反转录合成病毒RNA的双链cDNA,TaqⅠ酶切双链cDNA,加接头分子,并以接头分子为引物非特异扩增病原基因;PCR产物克隆后,酶切鉴定并测序,经GenBankBLAST软件进行序列分析。结果非序列依赖的单引物扩增HIV患者血清中外源基因得到多个DNA片段;重组克隆质粒经EcoRⅠ和HindⅢ双酶切,可见插入片段存在;将所有PCR产物测序,序列与已知病原基因序列一致。结论应用DNase处理及非序列依赖的单引物扩增方法可以检测RNA病毒性病原体。     

N-二(4-乙基苯-二氨基甲酰基)甲基磷酸与稀土配合物的合成及其与DNA作用的电化学研究

合成了一种结构易控、化学稳定性好、呈电中性及电化学活性高的含磷三足配体N-二(4-乙基苯-二氨基甲酰基)甲基磷酸及其稀土Eu(Ⅲ)配合物,采用了~1HNMR、~(13)CNMR、红外光谱、元素分析、差热-热重及连续物质的量比法对其组成进行表征并推断其结构,确定了配体与稀土离子形成了1∶1型配合物Eu(pic)3L。在进行配合物与DNA相互作用实验中,考察了扫描速率ν、p H、配合物浓度和离子强度因素对其的影响。实验结果表明,Eu(pic)3L与DNA碱基进行相互结合从而形成非电活性物质,使游离的Eu(pic)3L浓度降低,Eu(pic)3L的峰电流减小,说明Eu(pic)3L与DNA的相互作用是以嵌插形式结合的。根据Eu(pic)3L的电化学活性研究了以该配合物为杂交探针在DNA电化学传感器方面的应用。结果发现,Eu(pic)3L在修饰单链DNA的电极检测作用时,并无明显的电化学信号响应;检测杂交双链DNA时,却出现了明显信号,与此同时Eu(pic)3L的DNA电化学传感器对DNA的互补、碱基错配及非互补序列均有良好的识别功能。     

Insect antifeedant properties of anthranoids from the genusVismia

Vismiones and ferruginins, representatives of a new class of lypophilic anthranoids from the genusVismia were found to inhibit feeding in larvae of species ofSpodoptera, Heliothis, and inLocusta migratoria.     

Direct observation of freeze-thaw instability of latex coatings via high pressure freezing and cryogenic SEM

Despite its industrial importance, the subject of freeze-thaw (F/T) stability of latex coatings has not been studied extensively. There is also a lack of fundamental understanding about the process and the mechanisms through which a coating becomes destabilized. High pressure (2100 bar) freezing fixes the state of water-suspended particles of polymer binder and inorganic pigments without the growth of ice crystals during freezing that produce artifacts in direct imaging scanning electron microscopy (SEM) of fracture surfaces of frozen coatings. We show that by incorporating copolymerizable functional monomers, it is possible to achieve F/T stability in polymer latexes and in low-VOC paints, as judged by the microstructures revealed by the cryogenic SEM technique. Particle coalescence as well as pigment segregation in F/T unstable systems are visualized. In order to achieve F/T stability in paints, latex particles must not flocculate and should provide protection to inorganic pigment and extender particles. Because of the unique capabilities of the cryogenic SEM, we are able to separate the effects of freezing and thawing, and study the influence of the rate of freezing and thawing on F/T stability. Destabilization can be caused by either freezing or thawing. A slow freezing process is more detrimental to F/T stability than a fast freezing process; the latter actually preserves suspension stability during freezing. Presented at the 82nd Annual Meeting of the Federation of Societies for Coatings Technology, October 27–29, 2004 in Chicago, IL. Tied for first place in The John A. Gordon Best Paper Competition.     

Ethanol and (−)-α-Pinene: Attractant Kairomones for Bark and Ambrosia Beetles in the Southeastern US

In 2002–2004, we examined the flight responses of 49 species of native and exotic bark and ambrosia beetles (Coleoptera: Scolytidae and Platypodidae) to traps baited with ethanol and/or (−)-α-pinene in the southeastern US. Eight field trials were conducted in mature pine stands in Alabama, Florida, Georgia, North Carolina, and South Carolina. Funnel traps baited with ethanol lures (release rate, about 0.6 g/day at 25–28°C) were attractive to ten species of ambrosia beetles (Ambrosiodmus tachygraphus, Anisandrus sayi, Dryoxylon onoharaensum, Monarthrum mali, Xyleborinus saxesenii, Xyleborus affinis, Xyleborus ferrugineus, Xylosandrus compactus, Xylosandrus crassiusculus, and Xylosandrus germanus) and two species of bark beetles (Cryptocarenus heveae and Hypothenemus sp.). Traps baited with (−)-α-pinene lures (release rate, 2–6 g/day at 25–28°C) were attractive to five bark beetle species (Dendroctonus terebrans, Hylastes porculus, Hylastes salebrosus, Hylastes tenuis, and Ips grandicollis) and one platypodid ambrosia beetle species (Myoplatypus flavicornis). Ethanol enhanced responses of some species (Xyleborus pubescens, H. porculus, H. salebrosus, H. tenuis, and Pityophthorus cariniceps) to traps baited with (−)-α-pinene in some locations. (−)-α-Pinene interrupted the response of some ambrosia beetle species to traps baited with ethanol, but only the response of D. onoharaensum was interrupted consistently at most locations. Of 23 species of ambrosia beetles captured in our field trials, nine were exotic and accounted for 70–97% of total catches of ambrosia beetles. Our results provide support for the continued use of separate traps baited with ethanol alone and ethanol with (−)-α-pinene to detect and monitor common bark and ambrosia beetles from the southeastern region of the US.     

Many Drugs of Abuse May Be Acutely Transformed to Dopamine,Norepinephrine and Epinephrine In Vivo

It is well established that a wide range of drugs of abuse acutely boost the signaling of the sympathetic nervous system and the hypothalamic–pituitary–adrenal (HPA) axis, where norepinephrine and epinephrine are major output molecules. This stimulatory effect is accompanied by such symptoms as elevated heart rate and blood pressure, more rapid breathing, increased body temperature and sweating, and pupillary dilation, as well as the intoxicating or euphoric subjective properties of the drug. While many drugs of abuse are thought to achieve their intoxicating effects by modulating the monoaminergic neurotransmitter systems (i.e., serotonin, norepinephrine, dopamine) by binding to these receptors or otherwise affecting their synaptic signaling, this paper puts forth the hypothesis that many of these drugs are actually acutely converted to catecholamines (dopamine, norepinephrine, epinephrine) in vivo, in addition to transformation to their known metabolites. In this manner, a range of stimulants, opioids, and psychedelics (as well as alcohol) may partially achieve their intoxicating properties, as well as side effects, due to this putative transformation to catecholamines. If this hypothesis is correct, it would alter our understanding of the basic biosynthetic pathways for generating these important signaling molecules, while also modifying our view of the neural substrates underlying substance abuse and dependence, including psychological stress-induced relapse. Importantly, there is a direct way to test the overarching hypothesis: administer (either centrally or peripherally) stable isotope versions of these drugs to model organisms such as rodents (or even to humans) and then use liquid chromatography-mass spectrometry to determine if the labeled drug is converted to labeled catecholamines in brain, blood plasma, or urine samples.     

Novel polyurethane coating technology through glycidyl carbamate chemistry

Glycidyl carbamate chemistry combines the excellent properties of polyurethanes with the crosslinking chemistry of epoxy resins. Glycidyl carbamate functional oligomers were synthesized by the reaction of polyfunctional isocyanate oligomers and glycidol. The oligomers were formulated into coatings with several amine functional crosslinkers at varying stoichiometric ratios and cured at different temperatures. Properties such as solvent resistance, hardness, and impact resistance were dependent on the composition and cure conditions. Most coatings had an excellent combination of properties. Studies were carried out to determine the kinetics of the curing reaction of the glycidyl carbamate functional oligomers with multifunctional and model amines. Detailed kinetic analysis of the curing reactions was also undertaken. The results indicated that the glycidyl carbamate functional group is more reactive than a glycidyl ether group. Presented at the 82nd Annual Meeting of the Federation of Societies for Coatings Technology, on October 27–29, 2004, in Chicago, IL.