基于结晶质量增大效应的石英晶体微天平检测技术研究

发布时间：2018-05-22 18:35

本文选题：原位结晶 + 结晶调控　；参考：《山东师范大学》2017年硕士论文

【摘要】：为了提高石英晶体微天平对生物活性分子的灵敏度,建立了基于石英晶体微天平的原位选择性结晶方法。在之前的很多研究中,都对自组装膜进行了科学的论述以及研究,自组装膜的调控作用具有成熟的理论依据,并在实际应用中展现了良好的性能。在我们的试验中所建立的自组装膜主要是通过具有不同调控作用的官能团的暴露情况实现对碳酸钙原位结晶的调控作用。在我们建立的信号放大机制中,选用的两种功能基团分别为-N(CH3)3和-COOH,其中-N(CH3)3作为碳酸钙在表面的结晶抑制剂,-COOH作为碳酸钙在表面的结晶诱导剂,在这两种功能基团的共同调配下,实现了对碳酸钙在石英晶体微天平晶片表面原位结晶的调控,为我们的方法在信号放大方面提供了可能性。我们对其他常用的检测肿瘤标志物的方法以及应用石英晶体微天平的信号放大技术做了一定的学习与比较。相比之下,我们建立的基于石英晶体微天平的原位选择性结晶方法具有原理简单、不依赖生物码、灵敏度高和选择性高的优点,在癌症的早期诊断方面展现了了良好的发展前景。本文的主要内容如下:1.碳酸钙晶体在金片上的表面结晶过程可以通过石英晶体微天平检测,通过调节两种不同功能基团的用量比例可以实现晶体生长的定量控制。本实验中碳酸钙原位选择性结晶过程用到的两种功能基团分别为-N(CH3)3和-COOH,获得的晶体具有均匀的尺寸大小,相同的形态及晶型。实验发现随着-COOH组分比例的增加,晶体的量呈现一定规律的增加趋势。实验证明对金表面晶体生长的定量控制可以作为一个有效的质量扩增机制,该扩增机制可以用于石英晶体微天平的信号放大,以此可以提高石英晶体微天平的分析灵敏度。2.将上述基于碳酸钙原位选择性结晶技术应用到QCM对DNA分子的检测中,结果发现,该方法将检测DNA信号扩大了1000多倍,提高了DNA检测的灵敏度。在检测DNA时,最低检测限LOD达到了2aM,并且线性检测范围宽,为10aM到1nM。该方法同时具有很高的选择性,可以区分识别完全配对的DNA与1个碱基错配的DNA,可应用于单核苷酸的多态性(SNP)检测。3.基于碳酸钙原位结晶的石英晶体微天平信号放大技术,在检测DNA时展现了良好的性能。为了扩大其检测领域,我们将其用于细胞的检测,本实验选用的是Ramos细胞。在引入适配体后,结合碳酸钙原位结晶的石英晶体微天平信号放大技术检测DNA的方法,实现Ramos细胞的检测,其最低检测限LOD为5个/ml,检测范围为5个/ml到6000个/ml。最后,我们将该方法与其他基于质量信号放大的石英晶体微天平检测方法进行了对比,通过对比,证明了我们的实验具有很好的灵敏度。4.我们对碳酸钙原位结晶的石英晶体微天平信号放大技术进行了总结。该方法为石英晶体微天平技术的发展提供了新视角、新思路,基于该方法,可建立各种生物活性分子的通用检测平台,相信在研究者们的进一步研究下,该方法可以真正的应用于临床,为癌症的诊疗提供新技术。
[Abstract]:In order to improve the sensitivity of quartz crystal microbalance to bioactive molecules, an in-situ selective crystallization method based on quartz crystal microbalance was established. In many previous studies, the self assembled monolayers were scientifically discussed and studied. The regulation of self assembled films has a mature theoretical basis and is shown in practical applications. In our experiment, the self assembled monolayers are mainly regulated by the exposure of functional groups with different regulatory functions. In the signal amplification mechanism we have established, two functional groups are selected as -N (CH3) 3 and -COOH, respectively, of which -N (CH3) 3 is used as calcium carbonate. As a crystalline inhibitor on the surface, -COOH is a crystal inducer on the surface of calcium carbonate. Under the joint deployment of these two functional groups, the regulation of the in-situ crystallization of calcium carbonate on the surface of the quartz crystal microbalance wafer is realized. It provides a possibility for our method to amplify the signal. We have detected other commonly used tumor markers. In contrast, the in situ selective crystallization method based on quartz crystal microbalance has the advantages of simple principle, no dependence on biological code, high sensitivity and high selectivity, and is shown in the early diagnosis of cancer. The main contents of this paper are as follows: 1. the surface crystallization process of calcium carbonate crystal on the gold plate can be detected by quartz crystal microbalance, and the quantitative control of crystal growth can be realized by adjusting the proportion of two different functional groups. In this experiment, two kinds of work used in the process of selective crystallization of calcium carbonate are used in this experiment. The energy groups are -N (CH3) 3 and -COOH respectively. The obtained crystals have uniform size, the same morphology and crystal shape. The experimental results show that the amount of crystal increases with the increase of the proportion of -COOH components. The experimental results show that the quantitative control of the crystal growth of the gold surface can be used as an effective mass amplification mechanism. The increase mechanism can be used to amplify the signal of quartz crystal microbalance, so that the sensitivity.2. of quartz crystal microbalance can be improved by applying the CaCO3 in situ selective crystallization technology to the detection of DNA molecules by QCM. The results show that the method will expand the detection of DNA signals by more than 1000 times and improve the sensitivity of DNA detection. When detecting DNA, the minimum detection limit of LOD reaches 2aM and has a wide range of linear detection range, which has high selectivity for 10aM to 1nM.. It can distinguish completely paired DNA from 1 base mismatch DNA, and can be applied to single nucleotide polymorphisms (SNP) to detect quartz crystal microbalance signal based on calcium carbonate in situ crystallization. In order to expand its detection field, we use it to detect the area of DNA. In order to expand its detection field, we use it for the detection of cells. This experiment selects Ramos cells. The method of detecting DNA with the quartz crystal microbalance signal amplification technique combined with the in situ crystallization of calcium carbonate is used to detect the Ramos cells, and the lowest detection is made. The detection limit of LOD is 5 /ml, and the detection range is 5 /ml to 6000 /ml.. We compare the method with other quartz crystal microbalance detection methods based on the mass signal amplification. By contrast, we have proved that our experiment has a good sensitivity.4., we have the quartz crystal microbalance signal amplification by the calcium carbonate in situ crystallization. This method provides a new perspective and new idea for the development of quartz crystal microbalance. Based on this method, a general detection platform for various bioactive molecules can be established. It is believed that under the further research of the researchers, this method can be truly applied to the clinic and provide new techniques for the diagnosis and treatment of cancer.
【学位授予单位】：山东师范大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：O799

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