Wu Yafeng, Chen Chengliang, Liu Songqin
School of Chemistry and Chemical Engineering, Southeast University, Nanjing, 210096, People's Republic of China.
Anal Chem. 2009 Feb 15;81(4):1600-7. doi: 10.1021/ac802345z.
A novel strategy for sensitive detection of biomarkers using horseradish peroxidase (HRP)-functionalized silica nanoparticles as the label is presented. The enzyme-functionalized silica nanoparticles were fabricated by coimmobilization of HRP and alpha-fetoprotein antibody (anti-AFP, the secondary antibody, Ab2), a model protein, onto the surface of SiO(2) nanoparticles using gamma-glycidoxypropyltrimethoxysilane (GPMS) as the linkage. Through "sandwiched" immunoreaction, the enzyme-functionalized silica nanoparticle labels were brought close to the surface of gold substrates, as confirmed by the scanning electron microscopy (SEM) images. Enhanced detection sensitivity was achieved where the large surface area of SiO(2) nanoparticle carriers increased the amount of HRP bound per sandwiched immunoreaction. The electrochemical and chemiluminescence measurement showed 29.5- and 61-fold increases in detection signals, respectively, in comparison with the traditional sandwich immunoassay. The improved particle synthesis using a "seed-particle growth" route yielded particles of narrow size distribution, which allowed consistent loading of HRP and anti-AFP on each microsphere and ensured subsequent immunosensing possessed high sensitivity and reproducibility. This strategy was successfully demonstrated as a simple, cost-effective, specific, and potent method to detect AFP in practical samples.
本文提出了一种以辣根过氧化物酶（HRP）功能化的二氧化硅纳米颗粒作为标记物来灵敏检测生物标志物的新策略。通过使用γ-缩水甘油氧基丙基三甲氧基硅烷（GPMS）作为连接剂，将HRP和甲胎蛋白抗体（抗AFP，二抗，Ab2，一种模型蛋白）共固定在SiO₂纳米颗粒表面，制备了酶功能化的二氧化硅纳米颗粒。通过“夹心”免疫反应，酶功能化的二氧化硅纳米颗粒标记物被带到金基底表面附近，扫描电子显微镜（SEM）图像证实了这一点。由于SiO₂纳米颗粒载体的大表面积增加了每次夹心免疫反应中结合的HRP量，从而实现了增强的检测灵敏度。电化学和化学发光测量表明，与传统夹心免疫分析相比，检测信号分别增加了29.5倍和61倍。采用“种子颗粒生长”路线改进的颗粒合成方法产生了尺寸分布窄的颗粒，这使得HRP和抗AFP在每个微球上的负载一致，并确保后续免疫传感具有高灵敏度和可重复性。该策略被成功证明是一种在实际样品中检测AFP的简单、经济高效、特异且有效的方法。
