Surface plasmon resonance immunosensor using Au nanoparticle for detection of TNT

In order to develop the fully integrated portable surface plasmon resonance (SPR) system for detection of explosives, the amplification strategy of SPR signal was investigated. Indirect competitive inhibition method allowed the middle-sized SPR sensor to detect trinitrotoluene (TNT) at ppt level. However, this enhanced SPR signal was not high enough to detect TNT at ppt level by a miniaturized SPR sensor. Therefore, localized surface plasmon resonance (LSPR) effect using Au nanoparticle as further signal amplification approach was used. The amplification method of indirect competitive inhibition and LSPR were combined together for fabrication of the immunosurface using Au nanoparticle. TNT detectable range of this immunosurface was from 10 ppt (10 pg/ml) to 100 ppb (100 ng/ml), which was almost comparable to that without Au nanoparticle. The observed resonance angle change due to binding monoclonal TNT antibody (M-TNT Ab) with the immunosurface modified with Au nanoparticle was amplified to four times higher than that in absence of Au nanoparticle.     

血液中C反应蛋白的SPR传感检测方法研究

利用表面等离子体共振（SPR）生物传感技术,建立了一种可以快速准确分析血样中C反应蛋白的新方法。SPR传感器采用Kretschmann结构的角度连续扫描方式,使用2个步进电机分别驱动棱镜和光电检测器件转动进行单一样本的高精度分析。将C反应蛋白抗体修饰于敏感芯片表面,通过SPR传感器对血样中C反应蛋白浓度进行检测分析。利用SPR方法和传统免疫比浊法分别检测标准C反应蛋白样本和200份感染性疾病儿童患者（7～10岁,男112名,女88名）的血液样本,结果表明：SPR方法检测标准样本的线性变化区域更大。在患儿血样的检测中,尽管2种方法的结果基本相同,但是SPR检测速度更快,样本需求量更小、重复性更佳。这表明SPR生物传感分析方法在C反应蛋白检测中比传统方法更具优势,有望在临床检验分析中得以广泛应用。     

SPR biosensor by using E. coli outer membrane layer with autodisplayed Z-domains

In this work, the Z-domain of protein A with IgG-binding activity was expressed on the outer membrane (OM) of E. coli as a fusion protein of AIDA-1 by using “autodisplay technology”. The OM of E. coli with the autodisplayed Z-domains was isolated and then the OM with the Z-domains was layered on the gold surface of SPR biosensor in order to prepare the orientation-controlled antibody layer. The properties of the OM layer were analyzed by cyclic voltammetry, impedance spectroscopy and AFM. The immunoassay by using the OM layer with Z-domains showed that the limit of detection was improved as much as 50-folds by the orientation-control effect of the Z-domains. The sensitivity of assay (y-axis) at the each analyte concentration (x-axis) was also observed to be far improved by the orientation-control. The OM layer was then prepared on the SPR biosensor, and the far improved sensitivity from the orientation-control effect by the Z-domains was presented in comparison to the randomly oriented antibody layer by using hIgG as an analyte. For the regeneration of the SPR biosensor with the antibody layer immobilized to the Z-domains of OM layer, a cross-linker called glutaraldehyde was used for the covalent immobilization of the antibodies, and the feasibility of this method was demonstrated by performing repeated regeneration processes.     

Application of a functionalized parylene film as a linker layer of SPR biosensor

The surface plasmon resonance (SPR) biosensors have been used to detect various target analytes by using highly specific antigen-antibody interactions. In this work, a parylene film modified to have primary amine groups was applied as a linker layer of the SPR biosensor, and the primary amine groups were used for the covalent immobilization of proteins to the SPR biosensor. The feasibility of the parylene film as a linker layer was presented by estimating the influence of the parylene film on the SPR measurement parameters, such as the sensitivity and the detection range. Then, a model protein called horseradish peroxidase (HRP) was used to demonstrate the improved immobilization efficiency as well as the sensitivity of the SPR biosensor with the parylene-A film. Additionally, a reconstruction method of the immunoaffinity layer was presented by using oxygen plasma.     

Determination of 3-nitrotyrosine in human urine samples by surface plasmon resonance immunoassay

This paper describes the detection of a low-molecular weight molecule, 3-nitrotyrosine (3-NT) (∼226 Da), in human urine by coupling indirect inhibition assay with a surface plasmon resonance (SPR) sensor. 3-NT antibody (anti-3-NT Ab, mouse IgG) was used in this assay. An optimal antibody concentration has been measured at 27.9 μg/mL in order to obtain the best performance of the sensor surface. The lowest detection limit for 3-NT with this method is 4.7 ng/mL (S/N = 3). Sensor reliability was demonstrated by good specificity, intra-assay and inter-assay relative standard deviations <8%, average recovery of 107.68 ± 19.4% and sensor surface (self-assembled monolayer) stability through more than 200 regeneration cycles and 15 days of repeated measurement. This is the first SPR biosensor assay of 3-NT in human urine. The high stability of the SPR sensor surface underlies the potential of the SPR method as a low cost diagnostic tool for clinical detection of 3-NT.     

利用参比型SPR生物传感器进行SARS病毒抗原检测的研究

参比型SPR生物传感器是一种可以在单通道内实现对检测、参比两个位点同时检测的新型SPR分析系统.利用参比型SPR生物传感器在线制备了两种SARS病毒抗原的检测芯片.直接固定抗体法制备的检测芯片在检测时基本无反应;蛋白A固定抗体法制备的芯片在通入灭活SARS病毒时检测信号明显,参比信号略有上升.第二种芯片可以直接检测到病毒培养液上清中的灭活SARS病毒,检测灵敏度已达到1.66×104 PFU/mL.参比位点可以用来检测待测物中杂质引起的非特异性吸附.     

SPR生物传感器在急性白血病髓系抗原CD33检测中的应用

采用自组装膜技术,在传感芯片表面修饰抗CD33单克隆抗体,利用自行构建的SPR生物传感器检测人骨髓血液中急性白血病髓系抗原CD33的表达.实验结果表明SPR生物传感器对AML患者骨髓血液的响应值远远大于对健康人骨髓血液的响应值.与传统的流式细胞术(FCM)方法相比,SPR方法具有能快速给出定量分析结果,操作简单等优点....     

Surface plasmon resonance detection of small molecule using split aptamer fragments

It was difficult to detect small molecules directly using conventional surface plasmon resonance (SPR) biosensors since the changes of refractive index, which was resulted by binding small molecules, were usually small. In this paper, split aptamer fragments were used for the construction of SPR biosensor to determine small molecule such as adenosine with high sensitivity. An aptamer for adenosine was designed to be two flexible ssDNA pieces, one was tethered on Au film and the other was modified on Au nanoparticles (AuNPs). In the presence of adenosine, two ssDNA pieces reassembled into the intact aptamer structure and the AuNPs-labeled adenosine-aptamer complex was formed on the Au film. Then, the resonance wavelength shift was enhanced obviously, due to the electronic coupling between the localized plasmon of AuNPs and the surface plasmon wave associated with Au film. The results confirmed that this biosensor could detect adenosine with high sensitivity and selectivity. The limitation of detection (LOD) of this SPR biosensor was ca. 1.5 pM, which was an approximately ca. 2-3 order of magnitude lower than that of those SPR biosensors which utilized competitive methods.     

金纳米棒对SPR生物传感器灵敏度的增强效应

研究不同长径比的金纳米棒对表面等离子体共振(SPR)生物传感器灵敏度的增强效应.利用晶种生长法合成不同长径比的金纳米棒,并对其形貌、光学性质进行表征.采用双抗体夹心法,以金纳米棒 与羊抗人IgG的偶联体作为第二抗体,利用实验室自行研制的波长调制SPR生物传感器对人IgG进行测 试.实验结果表明:不同长径比的金纳米棒...     

动态范围可调的波导型SPR传感器模型

表面等离子体共振 (SPR)技术是一种简单直接的传感技术 ,SPR对金属表面附近的折射率的变化极为敏感 ,利用这一性质 ,表面等离子体共振传感器已成为生物传感器研究领域的热点。现提出一种电光调制波导型SPR模型 ,模拟计算表明该模型在不损害灵敏度的条件下 ,扩大了探测的动态范围。     

SPR生物传感器的应用现状与发展趋势

表面等离子体共振(SPR)生物传感器是一种基于物理光学原理的新型生化分析系统.与传统的超速离心、荧光法相比,具有实时检测、无需标记、耗样量少等特点.介绍了SPR生物传感器的基本原理,着重介绍了SPR生物传感器在生命科学,药物残留,疾病诊断以及食品检测中的应用,并对其发展趋势进行了展望.     

Enhanced surface plasmon resonance detection of DNA hybridization based on ZnO nanorod arrays

We demonstrated an enhanced surface plasmon resonance detection incorporating ZnO nanorod arrays (NRAs) built on a thin gold film. ZnO NRAs were fabricated by wet chemical growth method and used for the detection of DNA hybridization. Experimental results exhibited that ZnO NRAs provided a notable sensitivity improvement by more than 3 times, which is attributed to an increase in the surface reaction area. The measured sensitivity enhancement matched well with the numerical analyses based on the effective medium theory. Our approach is intended to show the feasibility and extend the applicability of the ZnO-based SPR biosensor to diverse biomolecular binding events.     

Ultrasensitive immunosensing of tuberculosis CFP-10 based on SPR spectroscopy

An antigen (Ag), CFP-10, found in tissue fluids of tuberculosis (TB) patients may be an ultimate candidate for use as a sensitive TB marker with a sensing method for early simplified diagnosis of TB. In this study, chemical and optical optimizations were carried out using novel immuno-materials for establishment of a self-assembled surface plasmon resonance (SPR) optical immunosensor system for detection of CFP-10, which is valuable for pre-clinical work, prior to conduct of massive clinical observations. For creation of a simple sensing interface, a monoclonal antibody (anti-CFP-10) was immobilized directly on a gold surface, followed by blocking with cystamine. Orientation and accessibility of anti-CFP-10 were assessed by the selective binding of CFP-10. Recent results indicate that the reusability of the sensor chip adopting the cystamine method was found to be preferable to other immobilization methods. A linear relationship was well correlated between SPR angle shift and CFP concentrations in the range from 100 ng mL⁻¹ to 1 μg mL⁻¹. Modification of the SPR chip with antibody provides a simple experimental platform for investigation of isolated proteins under experimental conditions resembling those of their native environment.     

Detection of sulphamethazine with an optical biosensor and anti-idiotypic antibodies

We describe the development of an assay for the detection of sulphamethazine in animal urine with a surface plasmon resonance (SPR) biosensor. In order to obtain a general assay that can easily be transferred to other veterinary drug residues, a monoclonal antibody against sulphamethazine and a corresponding anti-idiotypic antibody were used. The assay had a lower detection limit of 5 μg/l which is well below the maximum residue limits (MRL) of 100 μg/l and can be used for screening purposes in animal urine. The binding reaction between the antibodies as occurring during sensor application was characterized with respect to avidity and kinetic properties.     

利用图像SPR分析仪检测多组分蛋白芯片

介绍了一种新型高通量表面等离子体谐振(SPR)生化分析仪。基于图像分析技术和自动进样技术,该仪器可对微阵列SPR敏感芯片进行动态检测,实现高通量、多参数、多组分快速检测和定量分析。对兔IgG和人IgG分别与羊抗兔IgG和羊抗人IgG的免疫结合反应进行了实验检测。结果表明:该分析系统具有灵敏度高、免标记等优点,且阵列芯片中所设置的参比单元可以消除溶液本体折射率和温度变化的影响,提高了测量精度和准确性;另外,芯片可以再生重复使用,降低了测试成本。     

Development of a micro biochip integrated traveling wave micropumps and surface plasmon resonance imaging sensors

Since most of miniaturized surface plasmon resonance (SPR) sensing systems need commercially available peristaltic or syringe pumps, it is difficult to reduce the system size, biosample volume, and the production cost. In this paper, a compact biochip for clinical diagnosis is presented. The proposed biochip is integrated traveling wave micropumps and SPR imaging sensors on one chip. The micropump is composed of flexible microchannel and piezoelectric bimorph actuator array, and achieved the maximum flow rate 336 μl/min. The SPR imaging biosensor can quantitatively measure biosamples with multi microchannels, i.e. one biosample and two reference flows to obtain an analytical curve. The SPR imaging measurements with bovine serum albumin solutions were carried out using the prototype of the proposed diagnostic system composed of a pair of the micropump and the sensor. Since the clear SPR signal curve was observed, it was confirmed that the proposed system can be applicable to the clinical diagnosis.     

基于表面等离子体共振传感器的尿微量白蛋白检测

检测尿微量白蛋白(MAU)可对轻微肾脏损害进行早期诊断,也可对高血压患者可能发生的心脑血管事件进行预测。实验将抗体通过巯基自组装固定于Au膜表面,研制了一种快速检测MAU的表面等离子体共振(SPR)生物传感器。结果表明:直接检测法可以检测到0.3μg/L的MAU,而纳米Au放大法检测限可以低至0.03μg/L。     

SPR生物传感器在食品安全领域的应用研究

使用集成化手持式SpreetaTM SPR传感器快速检测大肠杆菌E.Coli 0157∶H7,利用大肠杆菌抗体的免疫吸附反应,采用亲和素—生物素系统放大检测的响应信号,并引入复合抗体作为第二抗体,整个检测过程在1 h内完成,实现病原微生物的快速检测。     

Highly sensitive surface plasmon resonance immunosensor for parts-per-trillion level detection of 2,4,6-trinitrophenol

A new surface plasmon resonance (SPR) immunosensor has been demonstrated for the determination of 2,4,6-trinitrophenol (TNP) based on the principle of indirect competitive immunoreaction using trinitrophenol–bovine serum albumin (TNP–BSA) conjugate and anti-TNP antibody. TNP–BSA conjugate was immobilized on a SPR chip by physical adsorption. TNP in solution competes with the immobilized TNP–BSA conjugate for binding with anti-TNP antibody, which inhibit the immunoreaction between TNP–BSA conjugate and anti-TNP antibody. The dependence of the inhibition to the concentration of TNP was utilized for quantification of TNP. Regeneration of the sensing surface for multiple analyses can be effected using pepsin solution. The sensor exhibited excellent sensitivity for the detection of TNP in a wide concentration range from 10 ppt to 100 ppb and has promising analytical characteristics, which can be extended to the determination of nitroaromatic explosive compounds for application to on-site detection of landmines.     

利用SPR生物传感器检测缺血修饰白蛋白

缺血修饰白蛋白(IMA)的检测对心肌缺血的早期诊断和治疗具有重要意义。利用纳米金颗粒和混合巯基自组装于金膜表面上,研制了一种快速检测IMA的表面等离子体共振(SPR)生物传感器。同时比较了直接法和抑制法的检测下限。结果表明:直接检测法可以检测到393ng/L的IMA,而抑制检测法检测限小于5.0 ng/L。与现有的IMA检测方法相比,SPR生物传感器具有特异性好、检测下限低以及检测耗时短等优点。