Fluorescence Turn-On Analysis of Trace Protein Based on Carbon Nanodots and Hybridization Chain Reaction

In this work, an ultrasensitive method for trace protein detection based on fluorescent carbon nanodots and hybridization chain reaction (HCR) is designed. Generally, the synthesized bright carbon nanodots are conjugated with two hairpin-structured DNA probes, respectively, which act as subsequent HCR fuel strands. Since single-stranded parts of DNA probes could be easily absorbed on graphene oxide (GO) nanosheets, fluorescence emission of carbon nanodots is effectively quenched via fluorescence resonance energy transfer. However, in the presence of target protein, the aptamer sequence in another hairpin-structured DNA probe specially interacts with target and the hairpin is opened. A single-stranded region is thus exposed, which initiates HCR by coupling with the DNA fuel strands on carbon nanodots. The formed HCR product displays a rigid, long double-stranded structure, which facilitates the release of carbon nanodots from GO surface. As a result, fluorescence of carbon nanodots is recovered and initial concentration of target protein can be estimated. This protein detection method shows a favorable linear response with a low limit of detection (2.3 fg mL⁻¹). Furthermore, it is highly selective and capable of detecting target in biological fluids like serum samples, which demonstrates the promising applications of this method.     

Mechanics of binding of a single integration-host-factor protein to DNA

We report on a single-molecule experiment where we directly observe local bending of a 76 base pair DNA oligomer caused by specific binding of a single integration-host-factor (IHF) protein. The conformational change of the DNA is detected by optically monitoring the displacement of a micron size bead tethered to a surface by the DNA. Since in the bound state the DNA loops around the IHF, a mechanical tension on the DNA tends to eject the protein. We measure how the rate for the protein to fall off the DNA depends on the mechanical tension in the DNA, gaining insight into the energy landscape for this molecular bond. Our method further demonstrates a new paradigm of molecular detection, where ligand binding is detected through the conformational change induced in the probe molecule. Here this allows the detection of single, unlabeled proteins.     

A robust infrared dim target detection method based on template filtering and saliency extraction

Dim target detection in infrared image with complex background and low signal-clutter ratio (SCR) is a significant and difficult task in the infrared target tracking system. A robust infrared dim target detection method based on template filtering and saliency extraction is proposed in this paper. The weighted gray map is obtained from the infrared image to highlight the target which is brighter than its neighbors and has weak correlation with its background. The target saliency map is then calculated by phase spectrum of Fourier Transform, so that the dim target detection could be converted to salient region extraction. The potential targets are finally extracted by combining the two maps. Moreover, position discrimination between targets in the two maps is used to exclude the false alarms and extract the targets. Experimental results on measured images indicate that our method is feasible, adaptable and robust in different backgrounds. The ROC (Receiver Operating Characteristic) curves obtained from the simulated images demonstrate the proposed method outperforms some existing typical methods in both detection rate and false alarm rate, for target detection with low SCR.     

Detection of PCR products amplified from DNA of epizootic pathogens using magnetic nanoparticles and SERS

Here we present a novel approach using surface‐enhanced Raman scattering (SERS) spectroscopy for the sequence‐specific detection of DNA utilizing magnetic nanoparticles (MNPs) for the enrichment of the target molecules. To achieve fast and efficient binding of longer DNA strands, e.g. PCR products, the hybridization procedure is performed in solution. To further purify and enrich the DNA strands of interest, MNPs are used for their separation. Following the binding of the target DNA, a dye‐modified, short synthetic ssDNA is hybridized, which serves as label for the SERS detection. The SERS spectra are used to identify the bound molecules. The applicability of this approach was first tested with short synthetic oligonucleotides to evaluate its specificity. Afterward, the system was applied to detect PCR products amplified from DNA of specific agents of epizootic diseases. Sequences of the bacterium Mycoplasma mycoides subspecies mycoides small colony type (MmmSC), causing contagious bovine pleuropneumonia (CBPP) were used as PCR targets. To demonstrate the multiplexing capability of SERS, the simultaneous detection of three different PCR products labeled with three dyes was performed. Copyright © 2010 John Wiley & Sons, Ltd.     

用光波导光模光谱技术研究DNA-DNA结合蛋白相互作用

探讨了应用光波导光模光谱(Optical waveguide lightmode spectroscopy,OWLS)技术研究DNA-DNA结合蛋白相互作用的可行性和灵敏性。以固定在传感器芯片表面的DNA探针为捕捉分子,溶液中同时含有探针结合序列和NF—κB结合位点序列的寡核苷酸与NF-κB亚单位p50同源二聚体形成的DNA-蛋白质复合物为检测分子,用光波导光模光谱检测技术建立非标记DNA-DNA结合蛋白相互作用检测研究体系。利用这一体系对不同样品中NF-κB p50浓度和具不同NF-κB结合位点序列的寡核苷酸与NF-κB p50亲合和力进行检测。样品中低至0．33 nmol/1的NF-κB p50被光波导光模光谱检测出,不同的NF-κB结合序列与NF-κB p50亲合力有显著的差异。研究发现,光波导光模光谱技术可以用于DNA-DNA结合蛋白相互作用研究,所建立的非标记检测研究体系可以进行样品中结合蛋白含量高灵敏检测和核酸序列与结合蛋白的亲合力的检测研究。     

Utilization of multiple phage display libraries for the identification of dissimilar peptide motifs that bind to a B7-1 monoclonal antibody

Summary Seven random peptide libraries (two displaying linear peptides and five displaying cysteine-constrained peptides) were constructed as gene III fusion proteins of the bacteriophage fd-tet. These libraries were used to screen a blocking monoclonal antibody raised against B7-1 (CD80), a human cell surface antigen that binds two T cell receptors, CD28 and CTLA-4. After three rounds of screening against the immobilized antibody, 1000-fold enrichment was observed in libraries displaying both linear and cysteineconstrained peptides. DNA sequencing of the enriched phage revealed two distinct consensus sequences: HXG(A/Y)XH and DVCXXGGPGC. Phage expressing these consensus sequences bound to L307.4 but not to an isotype matched antibody, indicating that binding was antibody specific. Synthetic peptides corresponding to both motifs inhibited phage binding to L307.4, indicating that the gene III protein is not required for peptide binding. In addition, the cyclized forms of synthetic peptides containing the DVCXXGGPGC motif were capable of inhibiting L307.4 binding to soluble B7-1/Fc fusion. Moreover, phage expressing only the HXG(A/Y)XH consensus sequence were inhibited from binding to L307.4 by the presence of chelating agents. These results indicate that the framework within which the peptide is presented on the surface of the phage may allow the identification of unique peptide motifs with distinct binding characteristics. These peptide motifs could be used for the design of peptidomimetics with therapeutic applications if they inhibit the binding of B7-1 to its T cell receptors.     

移位相加法日间探测低信噪比恒星

为实现白天红外光电测量系统对低信噪比恒星的质心及能量高精度计算,本文给出一种高效的方法。首先,分析了红外光学系统白天恒星的成像特征。其次,先对采集的图像序列进行预处理操作得到预处理图像。接着对预处理图像序列执行叠加求均值和下采样操作得到下采样图像。在下采样图像中以亮度为特征求取恒星的疑似位置后,在预处理图像序列上建立与疑似位置相对应的目标区域,在目标区域内顺序求取质心序列。对目标区域的图像序列以质心偏移为基础进行移位相加后获取目标图像。在目标图像上以信噪比为判据完成恒星提取,以及质心和能量的计算。再次,分析指出此方法能增强目标信噪比的原理,并给出其适用范围以及相关参数的确定方法。最后通过实验表明,采用移位相加法可增强目标的信噪比,并提高提取正确率;且对于SNR不大于4.8的恒星可将其质心和能量计算精度平均提高0.06 pixel和28.5%。移位相加法对低信噪比的恒星可较为精确地计算其质心和能量。     

Easy-to-implement method to target nonlinear systems

In this work we present a method to rapidly direct a chaotic system, to an aimed state or target, through a sequence of control perturbations, with few different amplitudes chosen according to the allowed control-parameter changes. We applied this procedure to the one-dimensional Logistic map, to the two-dimensional Henon map, and to the Double Scroll circuit described by a three-dimensional system of differential equations. Furthermore, for the Logistic map, we show numerically that the resulting trajectory (from the starting point to the target) goes along a stable manifold of the target. Moreover, using the Henon map, we create and stabilize unstable periodic orbits, and also verify the procedure robustness in the presence of noise. We apply our method to the Double Scroll circuit, without using any low-dimensional mapping to represent its dynamics, an improvement with respect to previous targeting methods only applied for experimental systems that are mapping-modeled. (c) 1998 American Institute of Physics.     

Specific heat spectra of non-interacting fermions in a quasiperiodic ladder sequence

We compute the specific heat spectra of non-interacting fermions whose energy spectrum was obtained from a quasiperiodic ladder sequence (Fibonacci and Rudin-Shapiro type), mimicking a DNA molecule model. The specific heat is calculated from their underlying multi-fractal energy spectrum, considering several values of energy densities. Comparisons are made with a real DNA sequence, namely the human chromosome 22 (Ch22).     

The application of magnetic force differentiation for the measurement of the affinity of peptide libraries

A new method has been developed for measuring the binding affinity of phage displayed peptides and a target protein using magnetic particles. The specific interaction between the phage displayed peptides and the target protein was subject to a force generated by the magnetic particle. The binding affinity was obtained by analyzing the force–bond lifetime.     

A Simple Model-Free Method for Direct Assessment of Fluorescent Ligand Binding by Linear Spectral Summation

Fluorescent tagged ligands are commonly used to determine binding to proteins. However, bound and free ligand concentrations are not directly determined. Instead the response in a fluorescent ligand titration experiment is considered to be proportional to the extent of binding and, therefore, the maximum value of binding is scaled to the total protein concentration. Here, a simple model-free method is presented to be performed in two steps. In the first step, normalized bound and free spectra of the ligand are determined. In the second step, these spectra are used to fit composite spectra as the sum of individual components or linear spectral summation. Using linear spectral summation, free and bound 1-Anilinonaphthalene-8-Sulfonic Acid (ANS) fluorescent ligand concentrations are directly calculated to determine ANS binding to tear lipocalin (TL), an archetypical ligand binding protein. Error analysis shows that the parameters that determine bound and free ligand concentrations were recovered with high certainty. The linear spectral summation method is feasible when fluorescence intensity is accompanied by a spectral shift upon protein binding. Computer simulations of the experiments of ANS binding to TL indicate that the method is feasible when the fluorescence spectral shift between bound and free forms of the ligand is just 8 nm. Ligands tagged with environmentally sensitive fluorescent dyes, e.g., dansyl chromophore, are particularly suitable for this method.

Improved gamut-constrained illuminant estimation by combining modified category correlation

In this paper we propose a new gamut-constrained illuminant estimation framework using an improved category correlation method. Firstly, we obtain a set of feasible illuminations by the original gamut mapping method. Then, the probability of each feasible illumination as the ground truth illuminant is calculated according to its ability to map the corrected image onto specific colors using the improved category correlation method. Differently from the original category correlation method, to decrease the effect of image noise and the computation complexity, instead of using an entire pixel set for estimating the probability of portable illuminant, superpixel segments of an input image are used in our improved method. And the best illuminant estimate is given on the basis of the measure of the degree to which each feasible illumination is consistent with the image data. Experiment results prove that our improved method shows better current state-of-the-art performance Gamut mapping methods.     

Bacillus subtilis转录因子CtsR蛋白中clpC操纵子结合区域的NMR研究

革兰氏阳性菌枯草芽孢杆菌（Bacillus subtilis）响应热刺激时,精氨酸激酶McsB磷酸化转录因子CtsR蛋白中clpC操纵子结合区域的Arg62（R）位点,使得CtsR与clpC操纵子解离,从而启动clpC相关基因的转录过程,以表达细菌应对热刺激所需的蛋白.本文以CtsR蛋白中结合clpC操纵子的区域（KRGGGG）为研究对象,通过对¹H NMR、¹H-¹H COSY、¹H-¹H TOCSY、¹H-¹⁵N HSQC和¹H-¹³C HSQC等谱图的综合分析,对其¹H、¹³C以及¹⁵N的化学位移进行归属,为该片段与clpC操纵子相互作用,以及精氨酸磷酸化调控机制的研究提供基础.