QCM免疫传感技术在宫颈癌变前检测的应用研究

发布时间：2018-01-27 23:00

本文关键词： QCM免疫传感技术复合谐振器质量灵敏度宫颈癌　出处：《电子科技大学》2016年博士论文　论文类型：学位论文

【摘要】：石英晶体微天平(Quartz Crystal Microbalance)是一种新型高灵敏度的质量传感器,由于其具有结构简单、成本低、检测速度快、测量精度可以达到纳克量级等优点,被广泛的应用于物理、化学、生物、医学等各个领域。QCM免疫传感器结合了QCM的高灵敏度和免疫反应的特异性,获得了研究人员的广泛关注,被广泛的应用于包括环境检测、食品安全、医学疾病诊断等各个领域。宫颈癌是所有女性疾病中致死率排名第二的恶性肿瘤,而在所有的恶性肿瘤中,宫颈癌也是最容易预防的恶性肿瘤。患者如果在宫颈癌的癌前病变时期得到了及时治疗,治愈的机会非常高,但该时期没有明显的临床症状,如果仅仅依靠临床表现无法对宫颈癌前病变进行诊断,而一旦宫颈癌前病变发展到浸润癌,病人一般会在2-5年的时间内死亡。因此,发展一种特异性强、灵敏度高的宫颈癌变前检测手段对宫颈癌的预防举足轻重。本文以压电免疫传感技术为基础,通过深入研究压电免疫传感器的检测机理,提出了一种宫颈癌变前检测的新方法。通过深入分析各种影响径向质量检测灵敏度的因素,搭建了适合宫颈癌变前检测的实验平台并设计了最适合该平台的实验方法,将该方法用于临床病人样本的检测,验证了该方法在宫颈癌变前检测中的合理性和有效性。本文的主要工作及创新点总结如下:(1)深入的分析了石英晶体微天平的检测机理,揭示了其具有高检测灵敏度的原因,同时,深入分析了石英晶体微天平表面振动位移分布情况,并以此为基础,详细研究了电极的半径、厚度及形状对石英晶体微天平质量检测灵敏度分布的影响,提出了提高质量检测灵敏度及改善质量检测灵敏度均匀性的方法,为石英晶体微天平的应用提供了理论基础。(2)研究了针对宫颈癌变前检测的特异性传感膜的制备方法。详细分析了各种特异性传感膜的制备方法,并通过实验从固化率、稳定性及免疫活性等各个方面对各种不同方法进行了比较,同时研究了环境温度、培育时间及抗体浓度等对传感膜固化率、稳定性及免疫活性的影响,最后提出了最适合用于宫颈癌变前检测的特异性传感膜制备方法及制备条件。(3)提出了一种基于QCM的宫颈癌变前检测的新方法。通过将该方法用于从四川大学华西附属第二妇女儿童医院获得的病人样本的检测发现,被测样本中P16~(INK4a)蛋白的含量和宫颈病变程度密切相关,病变程度越高,P16~(INK4a)蛋白含量越高,反之,病变程度越低,P16~(INK4a)蛋白含量就越少,并且在不同的病变程度之间,P16~(INK4a)蛋白的含量有明显的差异(p0.05),具有统计学意义。因此,通过检测样本中P16~(INK4a)蛋白的含量可确定被测对象的宫颈病变程度。通过将该方法的检测结果和病理医生的活检切片检查结果对比发现,该方法确定的病人病变程度和活检检测结果吻合度高。最后,本文还对该方法的特异性、稳定性、检测下限等性能进行了测试,为该方法的临床研究提供了基础。
[Abstract]:Quartz crystal microbalance (Quartz Crystal Microbalance) is a new kind of sensor with high sensitivity, because of its simple structure, low cost, fast detection speed, the measurement accuracy can reach nanogram level has been widely used in physics, chemistry, biology, medical field and other.QCM immunosensor combined with high specificity the sensitivity and the immune response of QCM, get the attention of many researchers, has been widely used in environmental detection, food safety, medical field disease diagnosis. Cervical cancer is the malignant tumor mortality rate ranked second of all female diseases, and in all the malignant tumors, cervical cancer is the most preventable the malignant tumor patients. If in the period of cervical cancer precancerous lesions got timely treatment, the chances are very high, but this time no obvious clinical symptoms, if only Based on clinical manifestations unable to diagnosis cervical precancerous lesions, and once the cervical precancerous lesions to the development of invasive carcinoma, patients often die within 2-5 years. Therefore, the development of a strong specificity and high sensitivity of cervical cancer prevention and detection methods of cervical cancer before the pivotal. On the basis of piezoelectric immunosensing technique, the detection mechanism of piezoelectric immunosensor through in-depth research, put forward a new method for the detection of cervical precancerous. Through in-depth analysis of various factors affecting the quality of radial detection sensitivity, to build an experimental platform for the detection of cervical cancer before change and design method is the most suitable for the platform, will the method for the detection of clinical samples of patients, the validation of the method in cervical cancer before the test is reasonable and effective. The main work and innovation points are summarized as follows: (1) in-depth analysis The detection mechanism of the quartz crystal microbalance, reveals its reason, high detection sensitivity at the same time, in-depth analysis of the surface displacement distribution of quartz crystal microbalance, and on this basis, a detailed study of the electrode radius, thickness and shape of quartz crystal micro balance quality detection sensitivity distribution, put forward to improve the quality of the detection sensitivity and improve the quality of detection sensitivity and uniformity of the method, and provides the theoretical basis for the application of quartz crystal microbalance. (2) studied the preparation method for the specific detection of cervical precancerous sensing film. A detailed analysis of the preparation of a variety of specific sensor membrane, and through the experiment from the curing rate the stability and immune activity, and other aspects of the comparison of different methods, and the effects of environment temperature, incubation time and antibody concentration on the sensing membrane curing rate, stability Effect and immune activity, and finally put forward the most suitable for cervical cancer before the specific detection sensing membrane preparation method and preparation conditions. (3) proposed a new method for the detection of cervical cancer QCM. Based on the detected by this method is used to obtain from the women and children hospital of Sichuan University affiliated Huaxi second the patient sample, the measured samples of P16~ (INK4a) is closely related to the protein content and the degree of cervical lesions, the higher the degree of lesion, P16~ (INK4a) protein content is high, on the contrary, the lower the degree of lesion, P16~ (INK4a) protein content was lower, and between the lesions in different degrees, (P16~ INK4a) have significant differences in protein content (P0.05), with statistical significance. Therefore, the detection of P16~ in the sample (INK4a) protein content can determine the degree of cervical lesions. The object of the detection results and the method of biopsy pathology doctor Compared with the results of biopsy, the results showed that the degree of coincidence between the patients determined by the method and the biopsy results was high. Finally, the specificity, stability and detection limit of the method were tested, which provided a basis for clinical research of this method.

【学位授予单位】：电子科技大学
【学位级别】：博士
【学位授予年份】：2016
【分类号】：TP212;R737.33

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