Two Simple NMR Experiments for Measuring Dipolar Couplings in Asparagine and Glutamine Side Chains

Residual dipolar couplings are now widely used for structure determination of biological macromolecules. Until recently, the main focus has been on measurement of dipolar couplings in the protein main chain. However, with the aim of more complete protein structure, it is also essential to have information on the orientation of protein side chains. In addition, residual dipolar couplings can potentially be employed to study molecular dynamics. In this Communication, two simple NH(2) and spin-state edited experiments are presented for rapid and convenient determination of five residual dipolar couplings from (15)N, (1)H correlation spectrum in asparagine and glutamine side chains. The pulse sequences are demonstrated on two proteins, 30.4-kDa Cel6A in diluted liquid crystal phase and 18-kDa human cardiac troponin C in water.     

Computational Mechanics of the Heart

Finite elasticity theory combined with finite element analysis provides the framework for analysing ventricular mechanics during the filling phase of the cardiac cycle, when cardiac cells are not actively contracting. The orthotropic properties of the passive tissue are described here by a “pole–zero” constitutive law, whose parameters are derived in part from a model of the underlying distributions of collagen fibres. These distributions are based on our observations of the fibrous-sheet laminar architecture of myocardial tissue. We illustrate the use of high order (cubic Hermite) basis functions in solving the Galerkin finite element stress equilibrium equations based on this orthotropic constitutive law and for incorporating the observed regional distributions of fibre and sheet orientations. Pressure–volume relations and 3D principal strains predicted by the model are compared with experimental observations. A model of active tissue properties, based on isolated muscle experiments, is also introduced in order to predict transmural distributions of 3D principal strains at the end of the contraction phase of the cardiac cycle. We end by offering a critique of the current model of ventricular mechanics and propose new challenges for future modellers. This revised version was published online in July 2006 with corrections to the Cover Date.     

Silicon nanowires as field-effect transducers for biosensor development: A review

The unique electronic properties and miniaturized dimensions of silicon nanowires (SiNWs) are attractive for label-free, real-time and sensitive detection of biomolecules. Sensors based on SiNWs operate as field effect transistors (FETs) and can be fabricated either by top–down or bottom–up approaches. Advances in fabrication methods have allowed for the control of physicochemical and electronic properties of SiNWs, providing opportunity for interfacing of SiNW-FET probes with intracellular environments. The Debye screening length is an important consideration that determines the performance and detection limits of SiNW-FET sensors, especially at physiologically relevant conditions of ionic strength (>100 mM). In this review, we discuss the construction and application of SiNW-FET sensors for detection of ions, nucleic acids and protein markers. Advantages and disadvantages of the top–down and bottom–up approaches for synthesis of SiNWs are discussed. An overview of various methods for surface functionalization of SiNWs for immobilization of selective chemistry is provided in the context of impact on the analytical performance of SiNW-FET sensors. In addition to in vitro examples, an overview of the progress of use of SiNW-FET sensors for ex vivo studies is also presented. This review concludes with a discussion of the future prospects of SiNW-FET sensors.     

Immobilization of insulin-like growth factor-1 onto thermosensitive hydrogels to enhance cardiac progenitor cell survival and differentiation under ischemic conditions

Stem cell therapy is a promising approach to treat myocardial infarction. However, direct delivery of stem cells into hearts experiences poor cell engraftment and differentiation, due to ischemic conditions (low nutrient and oxygen) in the infarct hearts. Development of suitable cell carriers capable of supporting cell survival and differentiation under these harsh conditions is critical for improving the efficacy of current stem cell therapy. In this work, we created a family of novel cell carriers based on thermosensitive hydrogels and insulin-like growth factor 1 (IGF-1), and investigated if these cell carriers can improve cell survival and differentiation under ischemic conditions. The thermosensitive hydrogels were synthesized from N-isopropylacrylamide, acrylic acid, acrylic acid N-hydroxysuccinicimide ester, and 2-hydroxyethyl methacrylate-oligo(hydroxybutyrate). The hydrogel solutions can be readily injected through 26G needles, and can quickly solidify at 37 °C to form highly flexible hydrogels. IGF-1 was immobilized into the hydrogels in order to support long-term cell survival and differentiation. Different amount of IGF-1 was immobilized by using hydrogels with different content of N-hydroxysuccinicimide ester groups. Cardiosphere derived cells were encapsulated in the hydrogels and cultured under ischemic conditions. The results demonstrated that a significant improvement of cell survival and differentiation was achieved after IGF-1 immobilization. These IGF-1 immobilized hydrogels have the potential to improve cell survival and differentiation in infarct hearts.     

多环芳烃暴露的生物标志物——尿中羟基多环芳烃

多环芳烃(PAHs) 是典型的持久性有机污染物,在职业高PAHs 暴露环境下,容易诱发肺癌、皮肤癌等癌症。对PAHs 的暴露评价可为流行病学研究和污染物风险评价等提供有效的数据。由于暴露途径的复杂化,采用尿样中PAHs 的代谢产物———羟基多环芳烃作为标志物来综合评价人体对PAHs 的内暴露情况已经成为研究的热点。本文系统介绍了多环芳烃的吸收、代谢、尿中PAHs 代谢产物的主要存在形式、主要的生物标志物以及它们的主要影响因素。     

Biological monitoring of 3-phenoxybenzoic acid in urine by an enzyme-linked immunosorbent assay

An enzyme-linked immunosorbent assay (ELISA) method was employed for determination of the pyrethroid biomarker, 3-phenoxybenzoic acid (3-PBA) in human urine samples. The optimized coating antigen concentration was 0.5 ng/mL with a dilution of 1:4000 for the 3-PBA antibody and 1:6000 for the enzyme conjugate. Urine samples were hydrolyzed with concentrated hydrochloric acid; extracted with dichloromethane and solvent-exchanged into a methanol/buffer solution, prior to analysis in a 96-microwell plate immunoassay. Quantitative recoveries of 3-PBA were obtained for fortified urine samples by ELISA (92 ± 18%) as well as by gas chromatography/mass spectrometry (GC/MS) (90 ± 13%). The overall method precision of these samples was within ±20% for both the ELISA and GC/MS methods. Analytical results from over one hundred urine samples showed that the ELISA and GC/MS data were highly correlated, with a correlation coefficient of 0.95. At the 10 ng/mL comparative concentration level, the false positive rate was 0% and the false negative rate was 0.8% for ELISA when using GC/MS as the reference method. The ELISA method has a suitable low detection limit for 3-PBA to assess pyrethroid exposures in non-occupational settings.     

基于铜掺杂碳纳米点荧光测定炭疽生物标志物

以柠檬酸、乙二胺为前体,硫酸铜为金属掺杂剂,采用一步水热法制备了一种高荧光量子产率的水溶性铜掺杂碳纳米点(Cu-CDs)。根据2,6-吡啶二甲酸(DPA)与碳纳米点的强螯合作用建立了铜掺杂碳纳米点荧光猝灭测定炭疽生物标志物DPA的新方法。在最佳实验条件下,DPA在5～100 nmol/L(r~2=0.994 1)和150～400 nmol/L(r~2=0.997 6)浓度范围内与Cu-CDs的荧光猝灭率呈良好的线性关系,检出限为2.3 nmol/L。该分析方法成本低、专属性强、灵敏度高、操作简便,在炭疽生物标志物检测方面具有良好的应用前景。     

2型糖尿病临床血清样本的内源性多肽定量组学分析

糖尿病是一种全身性代谢紊乱综合征，患者高血糖的形成与肝脏、胰脏、肠道、脂肪肌肉组织、肾脏、脑等多脏器的功能失调相关。在分子水平上呈现糖尿病的生物分子疾病谱图对糖尿病的临床早期诊断、分子分型及其病理过程的理解可提供更加全面的数据支持。本工作应用多肽组学分析技术，针对健康组、糖尿病前期组和2型糖尿病患者组临床血清样品进行内源性多肽定性、定量分析，共鉴定到690条可靠血清内源性肽段，其中163条为统计学差异血清内源性多肽，为2型糖尿病早期筛查、早期诊断及分子分型等提供了定量多肽组支持。     

A SPIN LABELING STUDY OF PIG CARDIAC TROPOMYOSIN

Tropomyosin (TM) extracted from pig cardiac muscle was spin-labeled with 2,2,6,6-tetramethyl-4-(dichlorotriazin)-aminopiperidine-1-oxyl. The ESR spectra of the product (SL-TM) were of a type of weak immobilization. Effects of three means for the denaturation were observed on the above spectra. The ESR spectrum obtained for SL-TMafter enzymatic degradation was found to be analogous to that for the label itself in a dilute solution and thereby the quantity of labels bound in SL-TM estimated. The Arrhenius plots attained through variable temperature measurement for SL-TM's exhibited two inflexion points (the conformational transition temperatures for TM) around 45℃ and 74-75℃, the latter temperature having not been reported in literature so far. However, the enzymatic degradation product from SL-TM behaved quite differently from it in the response to microwave power saturation and temperature variation.     

Current advances in electrochemical genosensors for detecting microRNA cancer markers

The electrochemical microRNA sensors are considered efficient, simple, and inexpensive analytical tools for the early diagnosis of cancer biomarkers. To enhance the sensitivity of the electrochemical genosensors toward detection of microRNAs, several amplification strategies based mainly on nanomaterials, enzymes, and oligonucleotides are investigated and discussed. This review highlights the main current achievements regarding the new promising and sensitive strategies for genosensors’ development, thus allowing for miroRNA analysis at the attomolar level.