Parallel detection and quantitative analysis of specific binding of proteins by oblique-incidence reflectivity difference technique in label-free format

In this work, we parallelly detected the specific binding between microarray targets including 12 different kinds of proteins and the probe solution containing five corresponding antibodies and quantitatively analyzed the interactions between CDH13 and solution phase anti-CDH13 at six different probe concentrations by oblique-incidence reflectivity difference (OIRD) method in label-free format. The detection sensitivity reached 10 ng/mL. The experimental results indicate that the OIRD method is a promising and competing technique not only in research work but also in clinic.     

Label-free high-throughput and real-time detections of protein interactions by oblique-incidence reflectivity difference method

Selected Mouse IgG of 1 mg/mL as target was fabricated on microarray for 500 sample dots.Label-free and real-time reaction dynamic processes were detected between the microarrays with Goat Anti-mouse IgG of 0.02 mg/mL using the obliqueincidence reflectivity difference(OIRD)method.We obtained the reaction results and the reaction dynamic curves of 500 protein dots.In addition,we also used label-free detection of protein microarrays of 10080 sample dots,including BSA and different concentrations of Mouse IgG and Rabbit IgG,by OIRD.The obtained reaction results between the protein microarray with 1 mg/mL Goat Anti-mouse IgG and 1 mg/mL Goat Anti-rabbit IgG are reported herein.Experimental results show that OIRD can be not only label-free high-throughput detection method for biological microarrays but also label-free real-time detection in the interaction processes of biomolecules.     

Detection of microarray protein biomolecules by oblique-incidence reflectivity difference technique without labelling agents

This paper reports that the detection to the protein in microarray format is carried out by oblique-incidence reflectivity difference （OI-RD） analysis without any labelling agents. The OI-RD intensities not only depend on the protein structure, but also vary with the protein concentration. The results indicate that this method should have potential application in detection of biochemical processes. The high throughout and in situ detection can be achieved by this method with further improving of the experimental system.     

用斜入射光反射差法无标记实时监测不同浓度兔IgG和山羊抗兔IgG反应的动力学过程

用斜入射光反射差法无标记实时监测1.25 mg/mL, 2.50 mg/mL和5.00 mg/mL三种不同浓度兔IgG和浓度为0.02 mg/mL山羊抗兔IgG反应的动态过程, 同时监测和获得三种不同浓度IgG反应的动力学曲线以及相应的反应时间. 实验结果表明, 斜入射光反射差法不仅能无标记地判别生物分子之间是否发生反应, 而且能无标记实时监测生物分子反应的动力学过程, 在生物分子相互作用的研究方面具有潜在和广泛的应用前景.     

Real-time and label-free detection of biomolecular interactions by oblique-incidence reflectivity difference method

We successfully conduct the label-free and real-time detection of the interactions between epoxy groups and rabbit IgG and 5' CTT CAG GTC ATG AGC CTG AT 3' oligonucleotide, and between the hybridization of 5' CTT CAG GTC ATG AGC CTG AT 3' and its complementary 3' GAA GTC CAG TAC TCG GAC TA 5' oligonucleotide, by the oblique-incidence reflectivity difference (OI-RD) method. The dynamic curves of OI-RD signals, corresponding to the kinetic processes of biomolecular combination or hybridization, are acquired. In our case, the combination of epoxy groups with rabbit IgG and 5' CTT CAG GTC ATG AGC CTG AT 3' oligonucleotide need almost one and a half hours and about two hundred seconds, respectively; and the hybridization of the two oligonucleotides needs about five hundred seconds. The experimental results show that the OI-RD is a promising method for the real-time and label-free detection of biomolecular interactions.     

Detection of hybridization of protein microarrays using an oblique-incidence reflectivity difference method

Mouse-Immunoglobulin G(mouse-IgG) with different concentrations in a range from 1000 to 0.0128 μg/mL and a specific hybridization with goat anti-mouse IgG were detected successfully by using an oblique-incidence reflectivity difference(OI-RD) method.Two detection signals,consisting of an imaginary part(Im{Δp-Δs}) and a real part(Re{Δp-Δs}) of OI-RD,were obtained simultaneously.The detection results of hybridization by OI-RD were in accord with that of traditional fluorescent scans.In particular,we label-freely detected the washed mouse-IgG microarray with a series of concentrations and acquired a linear correlation between OI-RD intensities and the protein concentrations in logarithmic coordinates.The detection sensitivity of OI-RD can reach 14 fg.These experimental results suggest that the OI-RD method has potential applications in proteomics and clinical diagnosis.     

Label-free detection of microarrays of biomolecules by oblique-incidence reflectivity difference microscopy

We developed an oblique-incidence reflectivity difference (OI-RD) scanning microscope for label-free imaging of microarrays of biomolecules upon solid substrates. We demonstrate that hybridization reactions in an oligonucleotide microarray fabricated upon a glass slide can be detected by such an OI-RD microscope.     

Evaluation of simulated reservoirs by using the oblique-incidence reflectivity difference technique

正Dear Editors,The evaluation of the oil-gas resources’potential depends on the characterization of reservoirs.The geometry of the pore-crack structures and the compositions of the pore walls have influence on hydrocarbon production about capillarity and permeability[1].Besides,it has been known that the adsorption phenomenon in micro-structure plays a significant role in oil-gas reservoirs.Some techniques have been applied to characterize conventional and unconventional reservoirs,     

Label-free detection of hybridization of oligonucleotides by oblique-incidence reflectivity difference method

The microarrays of 20-base oligonucleotide with different concentrations are detected before and after hybridization by the oblique-incidence reflectivity difference (OI-RD) method. The experimental results prove that OI-RD is a label-free method which can not only distinguish the concentration difference of oligonucleotides before and after the hybridization but also detect the hybridization of short oligonucleotides. At present the OI-RD method can detect 0.39 μmol/L 20-base oligonucleotide or less. These results suggest that the OI-RD method is a promising and potential technique for label-free detection of biological microarrays.     

Development of chemically modified glass surfaces for nucleic acid,protein and small molecule microarrays

Microarrays have become a widely used tool to investigate the living cell at different levels. DNA microarrays enable the expression analysis of thousand of genes simultaneously, while protein arrays investigate the properties and interactions of proteins with other proteins and with non-proteinaceous molecules. One crucial step in producing such microarrays is the permanent immobilization of samples on a solid surface. Our goal was to develop diverse linker systems capable of anchoring different biological samples, especially DNA and drug-like small molecules. We developed 6 different chemical surfaces having a 3-D-like linker system for biomolecule immobilization, and compared them to previously described immobilization strategies. The attachment chemistry utilizes the amino reactive properties of acrylic and epoxy functions. The capacity of the support was increased by creating a branching structure holding the reactive functions. The method of anchoring was investigated through a model reaction. From HPLC and mass spectrometry measurements we concluded that the covalent binding of DNA occurs through nucleobases. The tested systems offer the capability to permanently immobilize several biomolecular species in an array format.     

Description of surface roughness of sol-gel films/coatings by X-ray reflectivity technique

As an excellent optical or photoelectric material, indium tin oxide (ITO) film was prepared by sol-gel dip-coating and subsequent annealing process. X-ray reflectivity measurement based on the first Born approximation theory was performed for the characterization of surface roughness of ITO film. It is found that the roughness can be described as self-affined over finite length scales and the surface roughness increases with the annealing temperature or holding time. The results were compared with complementary data obtained by atomic force microscope tests and it is found that they match very well. The first Born approximation theory provides a valuable tool for the rough surface characterization of sol-gel films/coating through X-ray reflectivity technique.     

Measurement of inner surface roughness of capillary by an x-ray reflectivity method

The inner surface roughness of a capillary is investigated by the reflectivity of x-rays penetrating through the capillary. The results are consistent with the data from atomic force microscope (AFM). The roughness measured by this new method can reach the order of angstroms with high quality capillaries.     

X‐ray reflectivity measurements of liquid/solid interfaces under high hydrostatic pressure conditions

A high‐pressure cell for in situ X‐ray reflectivity measurements of liquid/solid interfaces at hydrostatic pressures up to 500 MPa (5 kbar), a pressure regime that is particularly important for the study of protein unfolding, is presented. The original set‐up of this hydrostatic high‐pressure cell is discussed and its unique properties are demonstrated by the investigation of pressure‐induced adsorption of the protein lysozyme onto hydrophobic silicon wafers. The presented results emphasize the enormous potential of X‐ray reflectivity studies under high hydrostatic pressure conditions for the in situ investigation of adsorption phenomena in biological systems.     

用光腔衰荡技术测量镜片的反射率

根据光腔衰荡光谱技术原理,建立了测量镜片反射率的实验装置。利用该装置测定了一对反射率相等的高反射腔镜,反射率测试结果为（99.925±0.001）%;以22.5°将直腔变为折叠腔,测得的反射率为（99.992±0.003）％。重复测定反射镜样品的反射率,精度达到10－5。该测量装置可用于超低损耗薄膜高反射镜反射率的精确测量。轻微移动探测器位置,对腔镜的测试结果影响不大。     

表面增强拉曼光谱技术检测新型冠状病毒刺突蛋白

开发具有高灵敏度、高准确性的新型冠状病毒（SARS-CoV-2）快速检测技术对疫情防控具有重要作用。本文利用表面增强拉曼光谱（SERS）技术对人体唾液中的痕量SARS-CoV-2病毒刺突蛋白（S蛋白）进行了检测。结果表明,含S蛋白的唾液样本与原始唾液样本的拉曼光谱具有显著区别,含S蛋白的唾液样本谱图中可清晰观察到属于S蛋白的拉曼谱线。该结果为后续SERS技术在SARS-CoV-2病毒快速检测方面的应用奠定了坚实基础。

     

Laser diagnostics of nanoscale dielectric films on absorbing substrate by differential reflectivity and ellipsometry

The differential reflection characteristics for ultrathin inhomogeneous dielectric film on absorbing substrate are investigated in the long-wavelength approximation. The obtained first-order expressions for differential reflectivity and changes in the ellipsometric angles caused by ultrathin layer are of immediate interest to the solution of the inverse problem. The method to determine the averaged values (not the realistic profile) of refractive index for inhomogeneous nanometric films are shown. The novel possibilities for determining the dielectric constant and thickness of nanoscale homogeneous films by the differential ellipsometric and reflectivity measurements are developed, and a simple method to estimate whether the nanometric film is homogeneous or not is also discussed.     

Laser diagnostics of nanoscale dielectric films on transparent substrate by integrating differential reflectivity and ellipsometry

The effect of nanometer dielectric films on the differential reflection characteristics of linearly polarized light from non-absorbing materials is investigated in the long-wavelength approximation. The second-order formulas for changes in the reflectance of s- and p-polarized light caused by ultrathin layer are obtained. A detailed analysis of the influence of ultrathin film to the reflectivity of p-polarized light in the vicinity of the Brewster angle is carried out. The novel methods are developed for determining the thickness and refractive index of uniform (or the average values of refractive index of nonuniform) nanometer-scale films by differential reflectivity and ellipsometric measurements.     

Nanometric thinning of bonded silicon wafers using sacrificial anodic oxidation and investigated by X-ray reflectivity

X-ray reflectivity and atomic force microscopy are used to investigate silicon oxide ultra-thin films. Quantitative results are shown using a reflectivity simulation model based on kinematical X-ray theory. Changes in film thickness are discussed in relation to current density, voltage, charge and anodization time. The density and resistivity of silicon oxide are calculated and compared to that of thermal oxide. The electrical field existing in the layer during anodization is estimated. Surface roughness is also measured locally and averaged over the entire surface, producing a low value that meets microelectronic requirements. Thickness is carefully controlled. We show that ultra-thin silicon oxide films are of very high quality. Similar investigations are made on a twisted bonded silicon substrate obtained by the molecular bonding of two silicon wafers. It is shown that the silicon oxide is also of very good quality and can be used as a sacrificial silicon oxide in thinning down the upper silicon film. Controlled, accurate thinning is achieved down to a thickness of 10 nm, the level which is required for etching the dislocation network present at the bonding interface.     

获得高精度频率差的关键技术

在利用LC振荡电路的参数变化而引起振荡频率的变化的特性进行的多种应用测量实验中,测量结果的精度都依赖于频率差Δf=f-f0.但当电路校准后进入测量状态时,电路参数会发生改变,使初始频率f0不复存在.如何在进入测量状态后保持初始频率f0以及精确提取频率差Δf,是提高测量精度的关键.为此提出一种采用锁相环自锁定技术和频率差计的方法,可以取得较好的效果.     

微晶硅薄膜的表面粗糙度及其生长机制的X射线掠角反射研究

采用等离子体增强化学气相沉积技术沉积一系列处于不同生长阶段的微晶硅薄膜.通过同步辐射X射线掠角反射技术研究微晶硅薄膜的表面粗糙度随时间等的演化，探讨微晶硅薄膜的生长动力学过程及其生长机制.研究结果表明，在衬底温度为200 ℃，电极间距为2 cm，沉积气压为6.66×1０² Pa，射频功率密度为0.22 W/cm²，氢稀释度分别为99%和98%的沉积条件下，在玻璃衬底上生长的微晶硅薄膜生长指数β分别为0.21±0.01和0.24±0.01.根据KPZ模型，微晶硅薄膜的生长机制为有限扩散生长. 关键词： X射线掠角反射 微晶硅薄膜 表面粗糙度 生长机制