School of Pharmacy, Nanjing University of Chinese Medicine, Nanjing, Jiangsu 210023, PR China.
School of Pharmacy, Nanjing University of Chinese Medicine, Nanjing, Jiangsu 210023, PR China.
Biosens Bioelectron. 2016 Nov 15;85:429-436. doi: 10.1016/j.bios.2016.05.039. Epub 2016 May 12.
An aptamer macroarray on a robust nanoplasmonic substrate with fluorescence enhancement is developed for a single-step sensitive detection of human platelet-derived growth factor-BB (PDGF-BB), a predominant cancer biomarker in cancer angiogenesis. A hybrid Au-nanoparticles-poly (dimethylsiloxane) (PDMS) as nanoplasmonic substrate is prepared via the in-situ reduction of AuCl4(-) ions in PDMS matrixes onto 96 or 384 well plates. In the absence of target molecules, unfolded PDGF-BB aptamer conjugated with dye TAMRA is electrostatically bound to a positively charged poly-L-lysine (PLL)-coated Au nanocomposites film surface, and the fluorescence enhancement effects can be optimized by varying the distance between TAMRA and the Au nanocomposites film, which is easily adjusted by varying the thickness of the biocompatible poly-(acrylic acid) (PAA/PLL) multilayers, and thus metal-enhanced fluorescence of dye TAMRA conjugated with the aptamer is generated up to 15.2-fold. The interaction of the aptamer to its target induces the reversible conformation change of the aptamer, and consequently, the electrostatic potential is overcome by binding force. As a result, the target-binding interaction of the aptamer causes the irreversible detachment of the aptamer from the nanostructured Au film surface to decrease fluorescence of TAMRA. The aptamer macroarray provides not only the appropriate sensitivity for clinical diagnostics with a wide range of linear detection from 10pg/mL to 10μg/mL, high specificity for PDGF-BB against VEGF-165, VEGF-121, NaCl and IgG, and temporal biological stability, but also a single-step detection. We envision that the efficient and robust aptamer macroarray can be extended to the detection of other biomarkers.
一种基于稳健纳米等离子体基底的适体微阵列，具有荧光增强功能，用于一步灵敏检测血小板衍生生长因子-BB（PDGF-BB），这是癌症血管生成中主要的癌症生物标志物。通过在 PDMS 基质中就地还原 AuCl4(-)离子，制备了 Au 纳米粒子-聚二甲基硅氧烷（PDMS）混合纳米等离子体基底，并将其应用于 96 或 384 孔板。在没有靶分子的情况下，未折叠的 PDGF-BB 适体与染料 TAMRA 缀合后通过静电作用结合到带正电荷的聚-L-赖氨酸（PLL）涂覆的 Au 纳米复合材料薄膜表面，通过改变 TAMRA 与 Au 纳米复合材料薄膜之间的距离，可以优化荧光增强效果，这可以通过改变生物相容性聚（丙烯酸）（PAA/PLL）多层的厚度来轻松调节，从而使与适体缀合的染料 TAMRA 产生高达 15.2 倍的金属增强荧光。适体与靶分子的相互作用诱导适体的可逆构象变化，从而通过结合力克服静电势。结果，适体与靶分子的结合相互作用导致适体从纳米结构的 Au 薄膜表面不可逆地脱离，从而降低 TAMRA 的荧光。适体微阵列不仅提供了广泛的线性检测范围（从 10pg/mL 到 10μg/mL）的临床诊断所需的适当灵敏度、对 PDGF-BB 相对于 VEGF-165、VEGF-121、NaCl 和 IgG 的高特异性以及时间生物学稳定性，而且还提供了一步检测。我们设想，这种高效稳健的适体微阵列可以扩展到其他生物标志物的检测。
