基于微流控芯片技术的表面增强拉曼研究

发布时间：2019-03-02 21:33

【摘要】：为了解决传统的检测方法存在的检测费事,费力,等待时间长等问题,实验采用了新的检测手段——将微流控芯片与表面增强拉曼结合的方法。即在一块微流控芯片上完成分析过程的样品反应、分离、检测等基本操作后,进行拉曼检测与分析,实验中选用纳米银粒子和纳米金球作为拉曼检测的基底,因金属纳米材料拥有独特的光电性质,特定尺寸的贵金属纳米粒子能够产生“热点”效应,SERS增强因子最高可达到107~1014倍,从而实现低浓度待测样品的高灵敏度的检测。本文的具体工作内容如下:(1)利用微流控滑动芯片作为检测载体,快速检测环境中和水产中的罗丹明6G和结晶紫含量。采用盐酸氢铵和硝酸银反应制得Ag NPs溶液,并将浓缩后的纳米银粒子和R6G分别加入滑动芯片中,选用多通道同时实验的方式,依靠微流控芯片的滑动将两者混合后,然后进行SERS检测,可以实现快速检测和样品的低消耗。实验发现在滑动芯片的检测区内浓度为10-10mol/L的罗丹明6G依然有明显的拉曼信号,在拉曼位移1360cm-1处呈现信号的峰值,并且随着反应混合时间的推移,其信号的强度也会有所变化,出现梯度最高值,浓度为10-10mol/L的最高信号强度为400,远远高于空白样品检测值。跟踪结晶紫(CV)进行相同实验,成功验证基于微流控滑动芯片的表面增强拉曼的高灵敏性检测。(2)提出利用回形针来制作三维微流控纸芯片的新方法,能够适应多种不同流体通道的微流控纸芯片。利用简单的比色检测来验证自主设计的纸芯片的流通性和反应灵敏性。在纸芯片上成功进行了牛血清白蛋白、二价铁离子的快速定量检测。牛血清白蛋白的线性范围是5-50μmol/L,检出限为0.15μmol/L,R2=0.982。二价铁离子的线性范围是0.6μmol/L-12μmol/L,检出限为0.18μmol/L,R2=0.992。验证了基于回形针的3D微流控纸芯片可用于实际样品检测的可能性。(3)采用SERS技术完成对纸芯片上肿瘤标志物CEA的检测。首先制备SERS探针标记CEA抗体,根据CEA抗原抗体的特异性结合作用,与连接在纸芯片上的固定抗体和抗原形成三明治‖免疫结构。利用SERS检测待测抗体上的MGITC信号,进而间接得到不同浓度抗原对应的信号值。CEA浓度分布在1 ng/m L-20ng/m L时,拉曼检测中标记分子的拉曼特征峰强度与检测物的浓度呈现出良好的线性关系,实现了对肿瘤蛋白标志物的高灵敏定量检测,CEA抗原的检出限为0.3ng/ml。这一检测方法经过拓展后可能适用于更复杂的多元肿瘤标志蛋白联合检测,具有广泛的应用前景。
[Abstract]:In order to solve the problems of detection cost, laborious and long waiting time existing in traditional detection methods, a new detection method, which combines microfluidic chip with surface-enhanced Raman method, is adopted in the experiment. After completing the basic operation of sample reaction, separation, detection and so on a microfluidic chip, Raman detection and analysis are carried out. In the experiment, nano-silver particles and gold nanoparticles are selected as the base of Raman detection. Due to the unique photoelectric properties of metal nanomaterials, the special size of precious metal nanoparticles can produce "hot spot" effect, and the SERS enhancement factor can be up to 107 脳 1014 times, thus realizing the high sensitivity detection of low concentration samples to be tested. The main contents of this paper are as follows: (1) the microfluidic slide chip is used as the detection carrier for rapid detection of Rhodamine 6G and Crystal Violet in environment and aquatic products. The Ag NPs solution was prepared by the reaction of ammonium hydrochlorid and silver nitrate. The concentrated silver nanoparticles and R6G were added to the slide chip respectively. The multi-channel simultaneous experiment was used to mix the two solutions by the sliding of the microfluidic chip. Then SERS detection can achieve rapid detection and low consumption of samples. It is found that Rhodamine 6G with the concentration of 10-10mol/L in the detection region of the sliding chip still has obvious Raman signal, and it shows the peak value of the signal at the Raman shift 1360cm-1, and with the passage of the reaction mixing time, the raman signal appears at the peak of the signal at the Raman shift 1360cm-1. The intensity of the signal also changes, and the maximum gradient value appears. The maximum signal intensity of 10-10mol/L is 400, which is much higher than that of blank sample. The high sensitivity detection of surface enhanced Raman based on microfluidic slide chip was successfully verified by the same experiment of tracking crystal violet (CV). (2) A new method of fabricating 3D microfluidic paper chip by using paper clip was proposed. Can adapt to a variety of different fluid channels microfluidic paper chip. A simple colorimetric test was used to verify the flowability and sensitivity of the self-designed paper chip. The rapid quantitative detection of bovine serum albumin (BSA) and divalent iron ion on paper chip was successfully carried out. The linear range of bovine serum albumin was 5-50 渭 mol / L, the detection limit was 0.15 渭 mol / L, and R2 was 0.982 2. The linear range of divalent iron ion is 0.6 渭 mol / L-12 渭 mol / L, the detection limit is 0.18 渭 mol / L, and R2 is 0.992.The detection limit is 0.18 渭 mol / L and the detection limit is 0.18 渭 mol / L. The possibility that 3D microfluidic paper chip based on paper clip can be used for real sample detection is verified. (3) the detection of tumor marker CEA on paper chip is completed by using SERS technology. First, the SERS probe labeled CEA antibody was prepared. According to the specific binding effect of CEA antigen antibody, the immobilization antibody and antigen formed the sandwich immune structure with the immobilized antibody and antigen attached to the paper chip. SERS was used to detect the MGITC signal on the antibody to be tested, and then the corresponding signal values of different concentrations of antigen were obtained indirectly. When the concentration of CEA was 1 ng/m L-20ng/m / L, There is a good linear relationship between the intensity of Raman characteristic peak of the labeled molecule and the concentration of the detection substance in Raman detection. The detection limit of CEA antigen is 0.3 ng / ml for the detection of tumor protein markers. This method may be applied to the combined detection of more complex multiple tumor marker proteins, and has a wide application prospect.
【学位授予单位】：烟台大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：O657.37

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