School of Chemistry and Chemical Engineering, Sun Yat-sen University , Guangzhou 510275, China.
Anal Chem. 2015 Nov 3;87(21):11039-47. doi: 10.1021/acs.analchem.5b03049. Epub 2015 Oct 13.
Poor selectivity and biocompability remain problems in applying surface-enhanced Raman spectroscopy (SERS) for direct detection of proteins due to similar spectra of most proteins and overlapping Raman bands in complex mixtures. To solve these problems, an aptamer recognition induced target-bridged strategy based on magnetic chitosan (MCS) and silver/chitosan nanoparticles (Ag@CS NPs) using SERS was developed for detection of protein benefiting from specific affinity of aptamers and biocompatibility of chitosan (CS). In this process, one aptamer (or antibody) modified MCS worked as capture probes through the affinity binding site of protein. The other aptamer modified Raman report molecules encapsulated Ag@CS NPs were used as SERS sensing probes based on the other binding site of protein. The sandwich complexes of aptamer (antibody)/protein/aptamer were separated easily with a magnet from biological samples, and the concentration of protein was indirectly reflected by the intensity variation of SERS signal of Raman report molecules. To explore the universality of the strategy, three different kinds of proteins including thrombin, platelet derived growth factor BB (PDGF BB) and immunoglobulin E (lgE) were investigated. The major advantages of this aptamer recognition induced target-bridged strategy are convenient operation with a magnet, stable signal expressing resulting from preventing loss of report molecules with the help of CS shell, and the avoidance of slow diffusion-limited kinetics problems occurring on a solid substrate. To demonstrate the feasibility of the proposed strategy, the method was applied to detection of PDGF BB in clinical samples. The limit of detection (LOD) of PDGF BB was estimated to be 3.2 pg/mL. The results obtained from human serum of healthy persons and cancer patients using the proposed strategy showed good agreement with that of the ELISA method but with wider linear range, more convenient operation, and lower cost. The proposed strategy holds great potential in highly sensitive and selective analysis of target proteins in complex biological samples.
由于大多数蛋白质的光谱相似且复杂混合物中的拉曼带重叠，因此在直接检测蛋白质时，表面增强拉曼光谱（SERS）的选择性和生物相容性仍然存在问题。为了解决这些问题，开发了一种基于磁性壳聚糖（MCS）和银/壳聚糖纳米粒子（Ag@CS NPs）的适体识别诱导靶桥接策略，用于检测蛋白质，该策略利用适体的特异性亲和力和壳聚糖（CS）的生物相容性。在这个过程中，一个修饰有适体（或抗体）的 MCS 通过蛋白质的亲和结合位点作为捕获探针。另一个修饰有拉曼报告分子的适体被封装在 Ag@CS NPs 中，作为基于蛋白质另一个结合位点的 SERS 传感探针。通过亲和结合，生物样品中的适体（抗体）/蛋白质/适体夹心复合物很容易被磁铁从生物样品中分离出来，并且蛋白质的浓度通过拉曼报告分子的 SERS 信号强度变化间接反映。为了探索该策略的通用性，研究了三种不同类型的蛋白质，包括凝血酶、血小板衍生生长因子 BB（PDGF BB）和免疫球蛋白 E（lgE）。该适体识别诱导靶桥接策略的主要优点是操作方便，具有磁铁，并且由于 CS 壳的帮助防止了报告分子的损失，从而信号稳定，避免了在固体基质上发生缓慢的扩散限制动力学问题。为了验证该策略的可行性，将该方法应用于临床样品中 PDGF BB 的检测。PDGF BB 的检测限（LOD）估计为 3.2 pg/mL。使用该策略对健康人和癌症患者的人血清进行检测的结果与 ELISA 方法的结果吻合良好，但具有更宽的线性范围、更方便的操作和更低的成本。该策略在复杂生物样品中目标蛋白质的高灵敏度和选择性分析方面具有很大的潜力。
