基于免疫层析术的光学生物传感技术研究
文献类型：学位论文
| 作者 | 黄立华 |
| 学位类别 | 博士 |
| 答辩日期 | 2009 |
| 授予单位 | 中国科学院上海光学精密机械研究所 |
| 导师 | 黄惠杰 |
| 关键词 | 光学生物传感器 免疫层析 上转换发光技术 Mie散射 纳米金颗粒 反射式光度计 |
| 其他题名 | Study of optical biosensor based on immunochromatography technology |
| 中文摘要 | 当今社会，人们在充分享受高速发展的科学技术造就的幸福生活的同时，也正遭受众多危险因素的威胁：快节奏的生活和高负荷的工作使得人们身体的抵抗力下降，极易受到病毒和细菌的侵袭；包括生物恐怖袭击在内的各种恐怖活动日益猖獗，严重威胁着世界和平和世界人民的生命安全；环境遭到严重破坏，且仍有恶化趋势，已侵害到人们的正常生活；食品中非法添加剂的使用和药物（农药与兽药）残留，使食品卫生与安全形势极为严峻，已对人们的健康造成重大伤害。所以，人们迫切需要快速、准确甄别上述威胁的检测方法。基于免疫层析术的光学生物传感器是满足这种需求的利器，它既保持了免疫层析术的敏感性、特异性和简便性，又具备了光学检测手段的灵敏、快速、无损等优点，已经并必将在医学诊断、环境监测、生物反恐、食品安全等领域发挥重要作用。本论文开展基于免疫层析术的光学生物传感器的理论与应用研究，具有重要的现实意义。 本论文的主要内容如下： 1) 全面综述了免疫层析型光学生物传感器的特点，着重论述了两种光学生物传感器——金标免疫层析型光学生物传感器和基于上转换发光技术（UPT）生物传感器的特点及国内外研究进展，指出了这两类仪器的发展趋势。 2) 结合理论推导与数值模拟分析了试纸条上免疫层析反应的微观过程。以双抗体夹心测抗原为例，采用微分方法求解了试纸条免疫层析的对流、扩散、反应三个方程，给出了近似解析解；进行了数值计算，得到了试纸条上免疫层析反应的动态过程；以上分析结果在第三章得到了实验验证。基于上述研究结果，详细阐述了免疫层析型光学生物传感器的总体设计、检测方式、控制和数据处理算法以及标定方法。 3) 详细阐述了金标免疫试纸条反射式光度计的工作原理与样机研制。详细讨论了纳米金颗粒的光散射和吸收特性，得出常用于免疫层析的纳米金颗粒的光吸收远大于光散射且对绿光吸收最为强烈的特性，由此建立了基于反射式光度计原理的金标免疫定量检测的方法及其理论模型。根据上述分析结果，研制出一台采用激光光源的金标免疫试纸条反射式光度计样机，实验结果如下：测量重复性优于2%；对hCG标准样品的检测灵敏度达到5mIU/ml，且至少在5mIU/ml~500mIU/ml范围内具有良好的线性响应特性（R2>0.99）；对cTnI标准样品的检测灵敏度达到1ng/ml，且在至少1ng/ml~50 ng/ml范围内具有良好的线性响应特性（R2>0.99）。 4) 在详细分析本实验室已研制成功的动态扫描型UPT生物传感器性能的基础上，提出了改进技术方案，包括采用照明光路与信号接收光路分离的光学系统、定时器控制电机运动的控制方式以及基于空间频谱分析滤波的功能带自动定位算法。改进后的UPT生物传感器可以自动准确地找到功能带（检测带与质控带）的位置，大大简化了操作步骤，且结构简单、装校方便。 5) 针对目前单重检测技术在对多种目标被检物进行检测时存在操作繁琐、效率低等不足，提出了一种多重生物检测技术，可一次检测一份样品中的多种目标被检物。详细阐述了一种基于上转换发光技术的多重检测试纸盘以及相应的多重检测光学生物传感器的设计，该仪器可在2分钟内实现对10种目标被检物的定量检测。 6) 总结了本论文取得的成果、创新点，以及进一步的研究内容。 |
| 英文摘要 | Today, along with the rapid development of science and technology, people can fully enjoy a happy life. Meanwhile, they are subjected to the threat of the many risk factors. Fast-paced life and high-intensity work impair immune function and decrease the body’s resistance to the virus and bacteria. A variety of terrorist activities, including bio-terrorist attacks, have become a serious threat to world peace and the safety of the people of the world. The seriously damaged environment has affected the people's normal life. Illegal additives in food and drug (pesticide and veterinary drug) residues hazard public health. It is the urgent need for rapid and sensitive detection method to respond to these threats. The method combined biology (immunochromatographic assay) and optics is very effective to meet the above demand. With the advantage of specificity of immunochromatography and rapidness of optical testing, the immunochromatographic optical biosensor have been widely used in many fields such as clinic diagnose, environmental monitor, anti-bioterrorism, food safety, etc.. This thesis focuses on the theory and application of optical biosensor with immunochromatographic assay. The primary contributions of this thesis are given in the following aspects: 1) The main characteristics of immunochromatographic optical biosensor are summarized. The emphasis was placed on the investigation of research progress of two kinds of optical biosensor, reflectance photometer based on gold-immune strip and UPT-based biosensor. Besides, the development trends were also presented. 2) The microscopic kinetics process of immunochromatography on test strip was investigated by theoretical analysis and numerical simulation. Take the double-antibody sandwich assay as an example, the convection-diffusion-reaction equations were solved with numerical differential method, and the evolution process of immunochromatography was obtained. Furthermore, the analytic solutions of complicated reaction process were presented. The above analysis result has been tested in experiments in the following chapter III. Based on the theoretical analysis and numerical simulation of the microscopic kinetics process, the design, test, control, data-processing algorithm and calibration method of immunochromatography optical biosensor has been elaborated. 3) The principle of reflectance photometer for gold labeled test strip was discussed and prototype was developed. The scattering and absorption behavior of nanogold particles were analyzed. The calculated results show that the absorption is much larger than the scattering and the intensive absorption of green light was observed. Based on the above calculated results, the prototype of reflectance photometer for gold labeled test strip was developed with laser as the source light. With the above reflectance photometer, serial dilutions of hCG were detected to achieve the test sensitivity of 5mIU/ml and the linear dynamic range of 5mIU/ml~500mIU/ml (R2>0.99). The test sensitivity of 1ng/ml and the linear dynamic range of 1ng/ml~50ng/ml (R2>0.99) were also obtained for cTnI dilutions. Besides, measurement repeatability is better than 2%. 4) Based on the developed scanning-type UPT bio-sensor, the improved technique program, such as the decoupled design of illumination and receiving optical system, control mode of motor with timer, and auto-searching algorithms of the boundaries of T line and C line with spatial spectrum analysis and the low-pass filter, were put forward. The improved UPT bio-sensor can search the boundaries of T line and C line automatically, which greatly simplifies the operation steps. Besides, the simple structure facilitates installation. 5) In order to overcome the disadvantage of cumbersome operation and low efficiency of single-anylte detection technology for multi-anylte detection, multi-anylte detection technology was put forward and several anyltes in one sample can be detected simultaneously. The design of multi-anylte detection bio-sensor was presented with upconverting phosphor particles as reporters. The quantitative measurement of up to ten analytes can be finished in two minutes. 6) In the end, the innovations of this thesis were summarized and some new research contents were put forward. |
| 语种 | 中文 |
| 源URL | [http://ir.siom.ac.cn/handle/181231/15403]  |
| 专题 | 上海光学精密机械研究所_学位论文 |
| 推荐引用方式 GB/T 7714 | 黄立华. 基于免疫层析术的光学生物传感技术研究[D]. 中国科学院上海光学精密机械研究所. 2009. |
入库方式： OAI收割
来源：上海光学精密机械研究所
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