Department of Applied Chemistry, College of Applied Science, Kyung Hee University, Yongin-si, Gyeonggi-do 446-701, South Korea.
Talanta. 2012 Sep 15;99:1030-4. doi: 10.1016/j.talanta.2012.08.002. Epub 2012 Aug 10.
We performed wide-range quantification of human thyroid-stimulating hormone (hTSH) using a gold nano-patterned sandwich immunoassay chip. Objective-type total internal reflection fluorescence microscopy (TIRFM) was used to detect hTSH at the single-molecule level. A gold spot with a diameter of 100 nm on a 10-mm square glass substrate was fabricated by electron beam nanolithography. When hTSH bound to antibodies conjugated to each 100-nm gold spot, there was an increase in the relative fluorescent intensity (RFI). The detection limit of this "TSH-nanoarray chip" was 360 zM (equivalent to five molecules), which demonstrated that a TSH-nanoarray chip could be used for detection at the single-molecule level. A linear response was observed over a wide dynamic range (from 360 zM to 36 pM, R=0.9812) without a fluorescence quenching effect. A significant enhancement in the sensitivity (~12,000-fold) was achieved with the 100-nm gold nano-patterned chip compared with results obtained using a traditional chemiluminescence immunoassay for the evaluation of TSH in human serum.
我们使用金纳米图案夹心免疫分析芯片对人促甲状腺激素（hTSH）进行了广泛的定量分析。采用基于全内反射荧光显微镜（TIRFM）的客观型检测方法，在单分子水平上检测 hTSH。通过电子束纳米光刻技术在 10mm 见方的玻璃衬底上制作了直径为 100nm 的金点。当 hTSH 与连接到每个 100nm 金点上的抗体结合时，相对荧光强度（RFI）会增加。该“TSH 纳米阵列芯片”的检测限为 360zM（相当于五个分子），表明 TSH 纳米阵列芯片可用于单分子水平的检测。在没有荧光猝灭效应的情况下，宽动态范围内（从 360zM 到 36pM，R=0.9812）观察到线性响应。与传统化学发光免疫分析法评估人血清中的 TSH 相比，使用 100nm 金纳米图案芯片可显著提高灵敏度（~12000 倍）。
