免疫磁珠荧光量子点技术检测结核分枝杆菌  

Detection of Mycobacterium tuberculosis using immunomagnetic beads fluorescence quantum dot method

作　　者：马慧[1,2] 王祎龙[3] 胡忠义[1] 秦莲花[1] 陈海珍[4] 陆俊梅[1] 王洁[1] 柏文娟[1] 郭琪[1] 高诗会[1] 杨华[1] 

机构地区：[1]同济大学附属上海市肺科医院结核病诊疗中心上海市结核病(肺)重点实验室,上海200433 [2]苏州大学基础医学与生物科学学院,苏州215123 [3]同济大学先进材料与纳米生物医学研究院,上海200092 [4]山西省儿童医院医学检验中心,太原030013

出　　处：《中国人兽共患病学报》2013年第5期466-471,共6页Chinese Journal of Zoonoses

基　　金：中央高校基本科研业务费专项基金(1511219013);上海市科委项目(No.10411955100,11nm0506100)联合资助~~

摘　　要：目的筛选结核分枝杆菌(MTB)免疫磁珠荧光量子点检测方法的最佳配体组合。方法将本室筛选获得的MTB结合肽H8和商业化抗MTB多克隆抗体(Pab)与纳米磁珠偶联,多肽H8、商业化抗MTB单克隆抗体(Mab)和抗热休克蛋白65单克隆抗体(Mabc)与荧光量子点偶联,分别获得2种免疫磁珠和三种功能化荧光量子点,两两组合用于MTB检测,通过显微镜下观察荧光信号以及激光诱导荧光仪测量荧光值,判断不同配体组合与MTB的亲和性及特异性,获得最佳组合及其检测下限。结果偶联Mabc的荧光量子点与免疫磁珠MNP-Pab组合检测MTB结果为阴性,其余5组均为阳性。测量荧光值,检测效果较好的两组组合分别是MNP-Pab+QD-H8和MNP-H8+QD-H8,阳性荧光值与阴性对照组比值分别是4.394和3.956,检测下限均是102条菌/mL。6组组合与12种非结核分枝杆菌(NTM)的亲和性各异,而与3种非分枝杆菌均不结合。结论 MNP-Pab+QD-H8和MNP-H8+QD-H8两组检测效果较好,与MTB亲和性较高,特异性较好,可作为MTB免疫磁珠荧光量子点检测技术的最佳配体组合。The aims of this study is to select the best ligand combinations of irnmunomagnetic beads and fluorescent quantum dots method for detecting Mycobacterium tuberculosis （MTB）. MTB-specific peptide H8 and commercial ami MTB polyclonal Pab were combined with magnetic nanoparticles, while the peptide H8 and two commercial anti-MTB monoclonal an- tibodies （Mab and Mabc） were coated with fluorescent quantum dots, to obtaine two kinds of immunomagnetic beads and tbree kinds of functionalized fluorescence qantum dot separately. The results show that anti MTB monoclonal antibodies Mabc coaled QDs combined with MNP Pab was negative, other five combinations detected MTB were all positive. The best two combina- tions were MNP-Pab＋QD-H8 and MNP-H8＋QD-H8, and the ratio of the fluorescence value between the experimental group and the control group were 4. 394 and 3. 956. Six groups, the limit detection of the two groups were all 10^2 CFU/mL. Six groups detected twelve kinds of nontuberculosis mycobacterium showed different affinity, and had not affinity with three kinds of non-mycobacterium. The combination of MNP Pab＋QD-H8 and MNP H8＋QD-H8 had better affinity and specificity, could be used as the best ligand combinations for immunomagnetic beads quantum dots method to detect MTB.

关 键 词：免疫磁珠 荧光量子点 分枝杆菌 结核 检测 配体 

分 类 号：R378[医药卫生—病原生物学]