量子点免疫荧光法在儿童非霍奇金淋巴瘤诊断中的应用

发布时间：2018-02-14 06:14

本文关键词： 非霍奇金淋巴瘤量子点免疫荧光法免疫酶组织化学快速特异　出处：《华中科技大学》2013年硕士论文　论文类型：学位论文

【摘要】：目的儿童淋巴瘤是小儿血液系统疾病中仅次于白血病的第二大恶性肿瘤，早期诊断对其治疗及预后起着非常重要的作用。量子点是近年来发展起来的一种新型荧光纳米微粒，具有激发光谱宽而连续、发射光谱窄而对称、荧光强度高、光化学稳定性好、耐光漂白等优点。本实验应用量子点(quantum dots, QDs)免疫荧光技术在石蜡包埋的非霍奇金淋巴瘤组织切片上同时检测两种抗原的表达，并与免疫酶组织化学方法相比较，探讨基于量子点诊断的快速特异性及优越性，以期扩展到其他病种的辅助诊断。方法（1）在温和的水相条件下制备出水溶性的巯基乙酸量子点，并利用不同发射波长量子点（525nm、605nm）标记的链霉亲和素复合物（QDs-SA）为基础的免疫荧光技术，分别检测T淋巴母细胞淋巴瘤、弥漫大B细胞淋巴瘤病例组织表面TDT、CD3和CD19、CD20两种抗原的共同表达；（2）利用免疫酶标技术在连续切片上依次标记上述抗原，比较两种方法的抗原定位效果，并评估量子点免疫荧光法的光学强度及稳定性。结果经修饰后的QDs-SA复合物与巯基乙酸量子点的光学特性变化不大，且实验中并未影响到组织细胞形态学特征。荧光显微镜下观察到几乎所有T淋巴母细胞淋巴瘤（17/20）都有TdT、CD3抗原的共同表达，，大多数弥漫大B细胞淋巴瘤（11/20）中也有CD19、CD20的共同表达，这与免疫酶组织化学技术检测结果定位完全一致，前者的阳性信号更强，有更高的光学稳定性，且无明显的荧光颗粒聚集现象，成像更清晰。结论表面修饰后的量子点有更好的组织相容性，并减少了其表面毒性。利用量子点免疫荧光法可以同时检测出组织石蜡切片中两种抗原的不同表达，这与免疫酶组织化学方法标记的抗原定位有高度一致性，相比较免疫酶标法一张切片仅标记一种抗原，可明显节约标本量，且有很高的阳性信号和强的光学稳定性。因此，量子点作为新型荧光探针，显示了其快速特异性及优越性，有望成为病理学诊断中的一种新的辅助检查手段。
[Abstract]:Objective Children's lymphoma is the second largest malignant tumor in children's hematological diseases, and early diagnosis plays an important role in its treatment and prognosis. Quantum dots (QDs) are a new type of fluorescent nanoparticles developed in recent years. The excitation spectrum is wide and continuous, the emission spectrum is narrow and symmetrical, the fluorescence intensity is high, the photochemical stability is good, In this experiment, quantum dot quantum dots (QDs) immunofluorescence technique was used to detect the expression of two kinds of antigens in paraffin-embedded non-Hodgkin 's lymphoma tissue sections, and compared with the immunohistochemical method. To explore the rapid specificity and superiority of quantum dot diagnosis (QDs) in order to extend to other auxiliary diagnosis. Methods 1) the water-soluble thioglycolic acid quantum dots were prepared under mild water phase conditions, and the immunofluorescence technique based on streptomycin complex QDs-SAwas used to label Streptomycin complex (QDs-SA) at different emission wavelengths. To detect the co-expression of TDTCD3 and CD19 CD20 on the tissue surface of T lymphoblastic lymphoma and diffuse large B cell lymphoma respectively. The antigen-localization effects of the two methods were compared, and the optical intensity and stability of the quantum dot immunofluorescence method were evaluated. Results the optical properties of modified QDs-SA complex and thioglycolic acid quantum dots changed little. Almost all T-lymphoblastic lymphomas (T-lymphoblastic lymphoma) were observed to have co-expression of TdTnCD3 antigen under fluorescence microscope, and CD19 / CD20 was also co-expressed in most diffuse large B-cell lymphomas (11 / 20). The positive signal of the former is stronger, the optical stability is higher, there is no obvious fluorescence particle aggregation, and the imaging is clearer. Conclusion the modified QDs have better histocompatibility and less surface toxicity. The different expression of two antigens in paraffin sections can be detected by using QDs immunofluorescence method. This is highly consistent with the localization of antigen labeled by immunohistochemical method. Compared with the immuno-labeled method, a section labeled with only one antigen can significantly save the sample size, and has a high positive signal and strong optical stability. As a new fluorescent probe, quantum dots (QDs) show its rapid specificity and superiority, which is expected to be a new auxiliary examination method in pathological diagnosis.
【学位授予单位】：华中科技大学
【学位级别】：硕士
【学位授予年份】：2013
【分类号】：R733.1

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