基于微流控传感器阵列的细胞免疫及结核感染同步检测芯片的研究

Tuberculosis is a common disease, which has high incidence, strong contagious and great harmful. Although Mycobacterium tuberculosis is the main pathogens, immune function of body and cell factors are closely correlated with tuberculosis incidence and disease outcome. So combined detection of cellular immune function and Mycobacterium tuberculosis infection is essential for clinical treatment and prognosis. Nowadays the flow cytometry and ELISA are respectively available, but there are some defects, for instance, expensive instrument, complicated operation, low the flux. The project will use micro fluidic, array sensing, immune capture and aptamer technology. Capture the T cells on different interdigital transducer through the antibody binding and count number of T cell. Then stimulate lymphocytes captured with tuberculosis -specific antigen to release IFN-γ、TNF-α, and perforin, which will be detect in situ with aptamers sensors detect. It would be implement synchronous detection of cell immune, cytokines and tuberculosis infection. The chip has many merits, for example, simple operation, high flux, fast speed, multi-target detection in one multi-functional etc.. This research will provide a new method for an early clinical diagnosis of tuberculosis infection, selecting the optimal treatment regimen, clarifying the occurrence of tuberculosis. It would promote the progress better and faster of the similar technologies and researches for clinic medicine.

结核病是临床常见病，其发病率高、传染性强、危害大，结核杆菌感染是其直接原因，但其发病、转归还与机体细胞免疫功能、细胞因子分泌密切相关，因此联合检测细胞免疫功能和结核感染对临床治疗和预后判断至关重要。目前，虽可用流式细胞术和ELISA等分别检测，但由仪器贵、操作繁、通量低等缺陷。本项目将微流控、阵列传感、免疫捕获及适体技术有机结合，将不同T细胞亚群由抗体捕获于不同叉指传感器，并完成细胞计数，遂以结核特异抗原刺激捕获的淋巴细胞，使之释放γ-干扰素、肿瘤坏死因子-α和穿孔素，用适配体修饰的传感器分别原位检测三种因子，以实现细胞免疫功能、细胞因子与结核感染的同步检测，该芯片具有操作简、特异性强、通量高、速度快等优点,集多目标多功能检测于一体。本研究将为临床及早诊断结核感染、选择最适治疗方案、阐明结核的发生发展规律提供新的技术手段，将推动同类技术和研究更好、更快地应用和服务于临床。

本项目基于微流控芯片、阵列传感器、免疫捕获及适配体技术，将不同T细胞亚群由抗体捕获于不同叉指传感器，并完成细胞计数，遂以结核特异抗原刺激捕获的淋巴细胞，使之释放γ-干扰素(IFN-r)、肿瘤坏死因子-α(TNF-α)和穿孔素(PFP)，用适配体修饰的传感器分别原位检测三种因子，以实现细胞免疫功能、细胞因子与结核感染的同步检测。构建出适宜微流控芯片集成组装的微纳传感器，并以此为基础设计出基于T淋巴细胞亚群计数、细胞因子分泌水平、结核菌感染同步检测的聚二甲基硅氧烷(PDMS)微流控芯片。2种细胞因子(IFN-r、TNF-α)应用现有适配体序列，1种(PFP)应用本项目筛选出的适配体序列，进行了纳米传感器的研究，构建出适宜微流控芯片集成的微纳传感器。对基于微流控传感器阵列的T淋巴细胞亚群计数进行了细胞捕获与细胞因子激活试验，取得了芯片构建的基础参数。结合碳纳米角和铂钯纳米线双重信号放大，构建了高灵敏度、适宜芯片集成的纳米传感器。构建了基于PDMS材料的数字芯片，正在进行该芯片在结核感染方面的试用性检测。本研究有望为结核病感染的诊断提供一种新的检测方法。