State Key Laboratory of Food Science and Technology, School of Food Science and Technology, Jiangnan University, Wuxi, Jiangsu, P R China.
Anal Chem. 2012 Jul 17;84(14):6263-70. doi: 10.1021/ac301534w. Epub 2012 Jul 6.
We presented a new aptasensor for mycotoxins, which was based on multiplexed fluorescence resonance energy transfer (FRET) between multicolor upconversion fluorescent nanoparticles (UCNPs) as donors and graphene oxide (GO) as the entire and effective acceptor. BaY(0.78)F(5):Yb(0.2), Er(0.02) and BaY(0.78)F(5):Yb(0.7), Tm(0.02) upconversion nanoparticles were synthesized and functionalized, respectively, with immobilized ochratoxin A (OTA)-aptamers and fumonisin B(1) (FB(1))-aptamers. On the basis of the strong π-π stacking effect between the nucleobases of the aptamers and the sp(2) atoms of GO, the aptamer modified-UCNPs can be brought in close proximity to the GO surface. The strong upconversion fluorescence both of BaY(0.78)F(5):Yb(0.2), Er(0.02) and BaY(0.78)F(5):Yb(0.2), Tm(0.02) can be completely quenched by the GO, because of a good overlap between the fluorescence emission of multicolor UCNPs and the absorption spectrum of GO. In contrast, in the presence of OTA and FB(1), the aptamers preferred to bind to their corresponding mycotoxins, which led to changes in the formation of aptamers, and therefore, aptamer modified-UCNPs were far away from the GO surface. Our study results showed that the fluorescence intensity of BaYF(5):Yb Er and BaYF(5):Yb Tm were related to the concentration of OTA and FB(1). We therefore developed a sensitive and simple platform for the simultaneous detection of OTA and FB(1) with multicolor UCNPs and GO as the FRET pair. The aptasensor provided a linear range from 0.05 to 100 ng·mL(-1) for OTA and 0.1 to 500 ng·mL(-1) for FB(1); the detection limit of OTA was 0.02 ng·mL(-1) and FB(1) was 0.1 ng·mL(-1). As a practical application, the aptasensor was used to monitor OTA and FB(1) level in naturally contaminated maize samples with the results consistent with that of a classic ELISA method. More importantly, the novel multiplexed FRET was established for the first time based on multiplexed energy donors to the entire energy acceptor; this work was expected to open up a new field of FRET system applications for various targets.
我们提出了一种新的真菌毒素适体传感器，它基于多色上转换荧光纳米粒子（UCNPs）作为供体与氧化石墨烯（GO）作为整体和有效受体之间的多重荧光共振能量转移（FRET）。分别合成并功能化 BaY(0.78)F(5):Yb(0.2)、Er(0.02)和 BaY(0.78)F(5):Yb(0.7)、Tm(0.02)上转换纳米粒子，分别固定化赭曲霉毒素 A（OTA）适体和伏马菌素 B1（FB1）适体。基于适体碱基与 GO 的 sp2 原子之间的强π-π堆积作用，修饰 UCNPs 的适体可以紧密接近 GO 表面。由于多色 UCNPs 的荧光发射与 GO 的吸收光谱之间存在良好的重叠，因此 BaY(0.78)F(5):Yb(0.2)、Er(0.02)和 BaY(0.78)F(5):Yb(0.2)、Tm(0.02)的强上转换荧光可以完全被 GO 猝灭。相比之下，在存在 OTA 和 FB1 的情况下，适体更倾向于与它们相应的真菌毒素结合，这导致适体的构象发生变化，因此，修饰 UCNPs 的适体远离 GO 表面。我们的研究结果表明，BaYF(5):Yb Er 和 BaYF(5):Yb Tm 的荧光强度与 OTA 和 FB1 的浓度有关。因此，我们使用多色 UCNPs 和 GO 作为 FRET 对开发了一种用于同时检测 OTA 和 FB1 的灵敏简单平台。适体传感器对 OTA 的线性范围为 0.05 至 100 ng·mL(-1)，对 FB1 的线性范围为 0.1 至 500 ng·mL(-1)；OTA 的检测限为 0.02 ng·mL(-1)，FB1 的检测限为 0.1 ng·mL(-1)。作为实际应用，该适体传感器用于监测天然污染玉米样品中的 OTA 和 FB1 水平，结果与经典 ELISA 方法一致。更重要的是，首次基于多能供体建立了新型多重 FRET，用于整体能量受体；这项工作有望为各种目标的 FRET 系统应用开辟一个新的领域。
