骨唾液蛋白纳米压电免疫传感器阵列的研制

Design of nano-piezoelectric immunosensor array to detect bone sialoprotein

目的以AT切型、10Hz双面镀金的石英晶体为换能器,结合纳米金颗粒独特的物理化学特性,制作一种用于临床血清骨唾液蛋白(bone sialoprotein,BSP)检测的纳米压电免疫传感器阵列。方法将纳米金颗粒通过1,6-己二硫醇固定在金电极表面,采用巯基化试剂Sulfo-LC-SPDP将鼠抗人BSP单克隆抗体巯基化,并将其固定于修饰有纳米金颗粒的石英晶体金电极表面,通过免疫反应检测正常人和骨质疏松患者血清中BSP的表达。结果 (1)对稀释度分别为1∶9,1∶8,1∶7,1∶6,1∶5,1∶4,1∶3,1∶2的参考标准血清进行检测,结果显示,在血清稀释度在1∶9～1∶2的范围内,构建的BSP纳米传感器阵列对血清检测的响应呈较好的线性关系,线性方程为y=872.66x-58.258,相关系数为0.974。(2)初步证明,骨质疏松症患者血清中BSP含量高于正常人血清,差异有统计学意义(t=20.162,P=0.000)。结论实验研制的BSP纳米压电免疫传感器阵列具有结构简单、操作容易、稳定性好、灵敏度高、特异性好等优点,可用于BSP的快速定量检测。

Objective With the unique properties of Au nanoparticles, a nano-piezoelectric immunosensor microarray was designed using AT-cut 10 Hz quartz crystal to detect blood sialoprotein （BSP） expression from clinical serum. Methods Monoclonal antibody of BSP was thiolated by heterobifunctional cross-linkers Sulfo-LC-SPDP. Next, the hydrosulfuryl-antibodies were fixed on the surface of gold-membrane electrode via Au nanoparticles modified to make a bio-sensitive membrane, which constructed the nano-piezoelectric immunity chips. BSP expression from the serum of healthy adults and patients with osteoporosis were examined after immunoreaction. Results The detection of reference standard serum at dilution of 1∶9, 1∶8, 1∶7, 1∶6, 1∶5, 1∶4, 1∶3 and 1∶2 showed that constructed BSP nano-immunosensor array had good linear relationship with serum detection results when the serum dilution ranged between 1∶9 and 1∶2; the linear equation was y = 872.66 x-58.258（correlation coefficient = 0.974）. The serum BSP expression levels of patients with osteoporosis were higher than those of healthy adults using BSP piezoelectric immunosensor array （t = 20.162, P = 0.000）. Conclusion The BSP nano-piezoelectric immunosensor array can be used for quantitative detection of serum BSP with the advantages of simple structure, easy operation, good stability, high sensitivity and specificity.