Detection of C-reactive protein on a functional poly(thiophene) self-assembled monolayer using surface plasmon resonance

The preparation of a new poly(thiophene) with pendant N-hydroxysuccinimide ester groups and its application to immobilization of biomolecules are reported. A thiophene derivative of N-hydroxysuccinimide ester was polymerized with FeCl(3) in chloroform and the resulting poly(thiophene) was characterized by nuclear magnetic resonance (NMR), Fourier transform infrared (FT-IR), and gel permeation chromatography (GPC). This polymer reacts with amine-bearing molecules to yield new poly(thiophene) derivatives and the specific interactions at the side groups could be detected. Thus, a self-assembled monolayer (SAM) using the polymer was formed on a gold-coated quartz cell and anti-C-reactive protein (anti-CRP) was immobilized. The binding behavior of CRP on the surface was monitored by use of a surface plasmon resonance (SPR) sensor system.     

角度指示型表面等离子共振传感器的研究

表面等离子体共振(SPR)技术被广泛应用于生物化学传感、药物分析和环境监测等领域。现基于表面等离子共振原理,在固定入射光波长为632.8nm的He-Ne激光入射下,采用镀有50nm银膜的Kretschmann棱镜型结构作为耦合器件,实现角度指示型SPR传感器对纯净水和酒精水溶液的检测。实验结果发现该SPR传感器具有较好的稳定性,其角度灵敏度达到219°/RIU。     

多孔分子印迹膜修饰的表面等离子体共振微囊藻毒素LR检测传感器

开发了一种多孔分子印迹膜修饰的表面等离子体共振(SPR)传感器,用于快速检测水中微囊藻毒素LR。研究了利用该传感器检测微囊藻毒素LR的方法。首先,通过原位聚合法在SPR传感芯片的裸金表面合成了微囊藻毒素LR的多孔分子印迹膜,制备出可以特异性捕获微囊藻毒素LR的SPR传感芯片。然后,利用Kretschmann棱镜耦合结构,构建了基于Kretschmann结构的波长调制型表面等离子体共振传感器。最后,通过检测不同浓度的微囊藻毒素LR溶液以及干扰物质微囊藻毒素RR溶液,研究了该传感器的测量范围、特异性等参数。结果表明,该传感器对于微囊藻毒素LR的检测灵敏度很高,可实现微囊藻毒素LR的定量检测,动态测量达2.1×10-9~1×10-6 mol/L。另外,传感器对于干扰物质微囊藻毒素RR无明显信号响应,表明传感器对于微囊藻毒素LR具有很好的特异性检测能力。     

Measuring the activation energy of thiol desorption using lateral force microscopy

Thermal stability of self-assembled monolayers (SAMs) is important for applications in various surface science applications. As a model material, 16-mercaptohexadecanoic acid (MHA) on template stripped gold surfaces was investigated to determine the effect of temperature on the change of lateral force signal using atomic force microscopy (AFM). Friction force signals were obtained at various temperatures in order to determine whether it was possible to correlate the friction signal with desorption of the thiol molecule from the surface. Samples were heated for up to 10 h ranging from 40 to 80 °C in air and scanned every hour. A kinetic model was introduced to correlate the lateral force signal to the activation energy of desorption of the SAM from gold surface with heating. The activation energy of the detachment using this technique is 25.4 kcal/mol, which is consistent with other more complex techniques.     

A simple method for the removal of thiols on gold surfaces using an NH4OH-H2O2-H2O solution

The removal behavior of self-assembled monolayers (SAMs) of thiol molecules on a gold substrate by an NH(4)OH-H(2)O(2)-H(2)O solution was studied using attenuated total reflectance infrared spectroscopy (ATR-IR) and atomic force microscopy (AFM). Furthermore, the impact of the concentration of NH(4)OH and H(2)O(2) in the solution and reaction temperature on the SAM removal rate and efficiency was explored. The SAM removal rate and efficiency were significantly influenced by the concentration of NH(4)OH rather than H(2)O(2). The solution containing the 2 : 1 molar ratio of NH(4)OH : H(2)O(2) among three different solutions showed the highest removal rate and efficiency in the removal of 11-mercapto-1-undecanol. The increase in the reaction temperature resulted in the enhancement on the SAM removal rate, but it led to the fast delamination of the gold layer. These results may be useful in the regeneration of sensor surfaces relying on gold/thiol chemistry.     

Chemical force microscopy of self-assembled monolayers on sputtered gold films patterned by phase separation

Patterned self-assembled monolayers (SAMs) were formed on gold films and observed by friction force microscopy (FFM) and adhesive force mapping with pulsed-force mode atomic force microscopy (PFM-AFM). The substrate gold films were prepared by sputtering gold on flat surfaces of osmium-coated cover glass with surface roughness, Ra, of 0.3 nm. The patterned samples with the CH3 and COOH terminated regions were prepared using the Langmuir-Blodgett (LB) method, partial removal of the LB film by ultrasonication, and SAM formation. The CH3 and COOH terminated regions of the patterned SAMs in air and in water were observed by mapping friction and adhesive forces with FFM and PFM-AFM, respectively, using gold-coated AFM tips chemically modified with a thiol compound terminating in CH3 or COOH. The adhesive forces measured in air increased in the order of CH3/CH3, CH3/COOH (or COOH/CH3) and COOH/COOH, while those in water increased in reverse order. The enormous high adhesive force observed in water for CH3/CH3 was attributed to hydrophobic interaction between the CH3 tip and the CH3 terminated sample surface. With CH3 tip, the lower friction force was observed, however, in water on the CH3 terminated region than on the COOH terminated region. This experimental finding raises a question as to what is the effective normal load in friction measurements in water.     

便携式SPR生化传感系统的研究

为了满足环境监测等多种场合的需要，设计了一种新型便携式表面等离子体谐振（Surface Plasmon Resonance，SPR）生化传感系统。系统包括驱动流通单元、SPR传感器单元和控制处理单元。驱动流通单元实现了两种样品自动进样检测；SPR传感器采用固定光路的结构，缩小了系统的体积；控制处理单元采用USB2.0接口芯片EZ-USB FX2系列作为数据传输的核心芯片，外接CPLD、RAM和高速AD，大大提高了系统的测量精度和灵敏度。     

AFM imaging in solution of protein-DNA complexes formed on DNA anchored to a gold surface

A chemical procedure for anchoring DNA molecules to gold surfaces was used to facilitate the imaging of DNA and DNA-protein complexes in buffer solution by tapping mode atomic force microscopy (TMAFM). For preparing flat gold surfaces, a novel approach was employed by evaporating small amounts of gold onto freshly cleaved mica to give flat films that were stable under aqueous buffer conditions. The thickness of the investigated films ranged from 1 to 10 nm. For typical films of 4-6 nm, which were stable under aqueous buffer conditions, the root mean square (RMS) roughness ranged between 0.25 and 0.5 nm, as measured by atomic force microscopy (AFM). This roughness is comparable to that of obtained by the template stripped gold (TSG) technique, which is widely used in scanning probe microscopy but involves more preparation steps. In order to visualize DNA and DNA-protein complexes by TMAFM, the DNA was chemisorbed to the gold surface through a linker carrying a terminal thiol group at the 5'-end of each of the DNA strands. The modified DNA fragments were bound to the gold films and imaged in buffer solution, while unmodified DNA could not be visualized. Since the DNA was not dried during the process, it can be assumed that its native conformation was retained. This mode of anchoring did not prevent interaction with proteins, as confirmed by the observation that the topology of a complex formed by adding the protein to a surface-anchored DNA was the same as that obtained by anchoring a pre-formed complex to the gold surface. We attribute this observation to the fact that the DNA is anchored to the gold surfaces only through its ends, therefore the DNA-support interaction is minimized but imaging is still possible.     

The immobilization of animal cells using the cysteine-modified RGD oligopeptide

RGD peptide sequence is an effective cell recognition motif and used to enhance the cell adhesion on desired solid material for cell immobilization. We have synthesized CRGD, CRGD-multiple-armed peptide (MAP), RGD-MAP-C and evaluated their comparative efficacy for cell immobilization. Each peptide was assembled on gold surface and investigated by the atomic force microscopy (AFM) technique in the contact mode. The viability of immobilized animal cells was examined by an MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay. Our results showed that RGD-MAP-C in comparison to others was the most effective proliferation of cells on the gold surface. The goal of this present work is integration to the nano-pattern cell chip bioplatform for biomedical assays or provide valuable insights into cell biology and design of biomaterials. This RGD-MAP-C can be applicable to the nano-pattern cell chip platform.     

基于Au-Ag复合膜的光纤表面等离子体共振传感器理论仿真研究

为增强基于Ag膜光纤表面等离子体共振(SPR)传感器的抗氧化能力,可将Au膜镀于Ag膜表面。利用TFCalc软件对不同厚度Ag膜和Au-Ag复合膜的光纤SPR传感特性进行理论仿真研究。仿真结果表明:光纤SPR吸收峰显著依赖于Ag膜厚度,当Ag膜厚度由40nm逐渐增加到80nm时,共振吸收峰的半峰全宽逐渐减小,且共振波长随Ag膜厚度的增大而减小,共振波长变化范围较小,仅为7nm左右;Au膜的引入对共振吸收峰反射率影响不大,不同厚度Au-Ag复合膜的SPR共振波长随Au膜厚度的增大而增大。     

Plasmonic Diagnostics for Tribology: In Situ Observations Using Surface Plasmon Resonance in Combination with Surface-Enhanced Raman Spectroscopy

The generation of transfer films is a common wear and lubrication mechanism of solid lubricants, such as polymers and lamellar solids. Material can transfer from a solid lubricant to a counter surface as early as the first cycle of sliding, initiating the formation of a transfer film, which can persist for the duration of sliding. Surface plasmon resonance (SPR) was used to monitor incipient molecular wear of a solid lubricant as performed here using a gold coated (50 nm) quartz prism in situ during sliding experiments. Surface-enhanced Raman spectroscopy (SERS) is a complementary technique enabling the analysis of ultra-thin transfer films. SPR and SERS experiments confirm that polytetrafluoroethylene and graphite transfer readily, with observed changes in SPR signal after one cycle of sliding, while ultra-high-molecular-weight polyethylene shows little transfer to the gold during sliding in the in situ SPR experiment. This shows the feasibility of SPR and SERS as important diagnostic tools for tribological studies.     

Fabrication of self-assembled oligophenylethynylenethiol monolayer for electrochemical glucose biosensor

An electrochemical glucose biosensor was developed using a gold (Au) electrode, which was composed of self-assembled oligophenylethynylenethiol monolayer and glucose oxidase (GOx) structure. Oligophenylethynylenethiol was used as a chemical linker for the immobilization of GOx on Au electrode, which facilitates the transfer of electron produced by enzyme reaction to the Au electrode. The electrical property of self-assembled oligophenylethynylenethiol monolayer was investigated by using cyclic voltammetry (CV). The formation of self-assembled oligophenylethynylenethiol monolayer and GOx layer on Au surface was verified by using atomic force microscopy (AFM) and surface plasmon resonance (SPR). The electrochemical glucose biosensor exhibited a linear relationship between target concentration and oxidation current in the range of 2–30 mM and its detection limit was 2 mM.     

Dynamic variations in adhesion of self-assembled monolayers on nanoasperities probed by atomic force microscopy

Octadecyltriethoxysilane (OTE) self- assembled monolayers (SAMs) and their effects on friction and adhesion have been investigated on various combinations of functionalized and unfunctionalized silicon oxide surfaces including the oxidized surface of crystalline Si(100), silica nanoparticle films, and oxidized Si atomic force microscopy (AFM) tips. Force-distance spectroscopy was utilized to probe and compare the properties of the OTE SAMs on silica asperities with nanoscale curvature against these same monolayers on surfaces with sub-1 nm roughness (flat surfaces). It was found that adhesion between SAMs and silicon oxide surfaces can vary significantly when assembly takes place on surfaces with nanoscopic curvature as compared to flat surfaces. Observations indicate that the SAM structure present during force measurements is dynamic in nature, which yields different adhesion values when measured with variations of both tip-sample contact time and tip-approach/retract rates. These results point the need in reporting a number of measurement parameters when probing adhesion by SAM functionalized tips.     

The effect of permanent dipole moments of adsorbates upon I-V characteristics of a bilayer tunneling junction between self-assembled monolayers on an Au(111) substrate and a gold tip

It is important to understand an electronic property of an interface between an organic material and a metal electrode. In the present work, we measured current-voltage (I-V) curves of self-assembled monolayers (SAMs) on Au(111) using a conducting atomic force microscope (AFM) with chemically modified Au-coated AFM tips. This contact resulted in a bilayer junction between the Au(111) substrate covered with one SAM and the Au-coated tip with the other SAM. An alkanethiol (octanethiol) and benzenemethanethiols with various terminal groups (-H, -CH(3), -Cl, -CF(3)) were used as the adsorbates. The shapes of the I-V curves depended on the terminal groups. This phenomenon was attributed to the change in the work function of gold due to different permanent dipole moments of the terminal groups.     

SPR生化分析仪及其在样品检测中的应用

表面等离子共振(SPR)是一种入射光照射到金属表面,导致金属表面产生等离子共振,反射光强出现衰减的现象,反射光强衰减程度与入射光波长、入射角及金属表面补测物质的折射率有关。根据这原理研制的SPR生化分析仪,在检测、分析分子间的相互作用等方面有着广泛的应用。针对HPSPR-6000生化分析仪的工作原理、技术参数、检测方法进行了详细的介绍,在芯片表面生长磺胺二甲嘧啶-BSA,并检测生长过程的响应信号,同时还对磺胺二甲嘧啶抗原与抗体在芯片表面的结合响应信号进行了检测。     

Tribological properties of asperity arrays coated with self-assembled monolayers

The effects of a self-assembled monolayer (SAM) coating on the friction and pull-off forces were determined by using two-dimensional asperity arrays on silicon wafers. The arrays were coated with SAM composed of one of five different alkylchlorsilanes. First, two-dimensional asperity arrays were created by using a focussed ion beam (FIB) system to mill patterns on silicon plates. Each silicon plate had different patterns of equally spaced asperities. Each pattern (5 × 5 μm²) had a different radius of curvature of the asperity peaks, ranging from about 200 to 2500 nm. Then, each silicon plate was immersed in a solution of a different alkylchlorsilane in hexane (either hexyltrichlorosilane, octyltrichlorosilane, dodecyltrichlorosilane, tetradecyltrichlorosilane, or octadecyltrichlorosilane), thus coating the asperity arrays with SAM. The friction and pull-off forces on the SAM-coated arrays were measured by using an atomic force microscope (AFM) that had a square flat probe. The pull-off force for SAM-coated silicon was roughly proportional to the radius of curvature of the asperity peaks. The magnitude of the pull-off force corresponded approximately to the capillary force calculated by using the contact angle of water on the surface of SAM. The friction coefficient correlated with the inverse of the alkyl-chain length of the SAM.     

Self-assembled monolayer on diamond-like carbon surface: formation and friction measurements

We have investigated self-assembled monolayers (SAMs) of heptadecafluoro-1,1,2,2-tetradecyltrietoxysilane (FTE) on diamond-like carbon (DLC) surfaces formed by a simple immersing process. SAM formation on DLC surfaces was verified by contact angle measurements, ellipsometry and X-ray photoelectron spectroscopy (XPS). Water and hexadecane contact angles increased gradually with immersing time and saturated at about 110 and 70 degrees, respectively. Ellipsometric measurements showed that the film thickness was 1.4 to 1.6 nm, which corresponded reasonably to the thickness of FTE monolayer. XPS data showed the presence of FTE molecules on the DLC surface. These results ensured the SAM formation of FTE molecules on the DLC surface.We further measured and compared the friction of unlubricated, SAM coated and 2 nm thick perfluoropolyether (PFPE) coated DLC surfaces using lateral force microscopy (LFM) as functions of the applied load and the sliding velocity. The SAM coated DLC surfaces showed lower friction than the unlubricated DLC surfaces and the friction coefficient decreased by about 15% compared to the unlubricated DLC surfaces. Scratch tests revealed that the critical load of the DLC film increased due to the SAM deposition. These results are attributed to the hydrophobic nature of the SAM coated surface. On the other hand, even though the water contact angle of the SAM coated surface was larger than the 2 nm thick PFPE coated surface, the friction of the SAM coated surface was larger than that of the PFPE coated surface. Also, the critical load of the SAM coated DLC surface in scratch test was lower than the PFPE coated surface. These results indicate that the hydrophobic nature of the surface is not the only factor which determines the friction characteristics in the nano-lubricating system, and it is attributed to the mobile characteristic of PFPE lubricant.     

基于纳米修饰透射式光纤传感测试

基于金属纳米颗粒结构的光学特性,结合光纤传感技术,对金属纳米颗粒的光纤传感特性进行了研究.实验中采用种子溶液生长法合成了粒径在60～80 nm的星形纳米金颗粒,以此作为光纤传感敏感部分的修饰体,修饰到锥形光纤表面作为表面拉曼增强基底.最后选取了不同浓度的酒精溶液对其进行了透射谱和拉曼谱测试,结果表明金属纳米颗粒的激发谱对周围介质特性非常敏感,同时对基于金属纳米颗粒锥形SERS基底的拉曼谱存在非常高的增强.     

Atomic force microscopy imaging of Xenopus laevis oocyte plasma membrane purified by ultracentrifugation

Atomic force microscopy (AFM) was used to investigate the native plasma membrane of Xenopus laevis (X. laevis) oocyte purified by means of ultracentrifugation on sucrose gradient and subsequently adsorbed on mica leaves through a physisorption process. Reproducible AFM topography images were collected, analyzed, and compared. AFM images showed the presence of large single or double bilayer membrane sheets covered with protein complexes. The lateral dimension and height of protein complexes imaged in air showed a normal distribution centred on 15.4 +/- 0.4 nm (mean +/- SE; n = 59) and 3.9 +/- 0.2 nm (mean +/- SE; n = 57), respectively. A density of about 270 protein complexes per square micron was calculated. Less frequently, ordered nanometer domains with densely packed protein complexes arranged in hexagonal patterns were also visualized in AFM images, confirming previously published data. Their lateral dimension and height showed a normal distribution centred on 23.0 +/- 0.4 nm (mean +/- SE; n = 42) and 1.5 +/- 0.6 nm (mean +/- SE; n = 90), respectively. A density of about 870 protein complexes per square micrometer was calculated. Advantages and drawbacks of this new sample preparation for AFM imaging are discussed.     

A study on conformational changes by electron charges in viologen single molecules by using STM

The topography of self-assembled viologen derivatives (VC(8)SH, VC(10)SH, HSC(8)VC(8)SH, and HSC(10)VC(10)SH) molecules on an octanethiol (C(8)) self-assembled monolayer (SAM) modified gold surface was measured using ultrahigh-vacuum scanning tunneling microscopy (UHV-STM). We demonstrate here a novel matrix SAM appropriate for isolation of the viologen molecules. The C(8) was used for a matrix SAM, in which the VC(8)SH, VC(10)SH, HSC(8)VC(8)SH, and HSC(10)VC(10)SH were inserted at molecular lattice defects. The isolated single molecules of viologen derivatives inserted in the matrix SAM were observed as protrusions in STM topography using a constant current mode. We measured the topographic heights (VC(8)SH: 1.53nm, VC(10)SH: 2.01nm, HSC(8)VC(8)SH: 2.71nm, and HSC(10)VC(10)SH: 3.3nm) of the molecular protrusions using STM. Also, changes in the central axis of viologen molecules were observed as VC(8)SH (0.5-0.73nm), VC(10)SH (0.4-0.74nm), HSC(8)VC(8)SH (0.67-0.84nm), and HSC(10)VC(10)SH (0.67-0.99nm), respectively.