组成图示
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传感器类型
荧光生物传感器
检测对象
甲胎蛋白（alpha-fetoprotein, AFP）；样品基质：磷酸盐缓冲液（PBS）和人血清（human serum）
检测原理
传感器以去包层PMMA光纤为换能基底，表面固定抗AFP捕获抗体并用BSA封闭。样品中的AFP与捕获抗体结合后，加入DyLight 649标记的抗AFP二抗-Au-PA探针，形成夹心复合物。658 nm激光在光纤中传播，去包层区产生倏逝波，激发Au-PA表面局域表面等离子体（LSP），在金纳米颗粒附近形成强局域电磁场，高效激发邻近DyLight 649荧光团，产生LSPCF/金属增强荧光。每个探针含多个荧光团同时被激发，实现信号放大。结合AFP越多，荧光信号越强，经680.4 nm窄带滤光片由PMT检测，锁相放大器提高信噪比，从而获得与AFP浓度对数相关的响应。
检测灵敏度
LOD: 0.1 ng/mL (1.4 pM) in PBS；血清最低可测: 2.33 ng/mL；线性范围: 0 ng/mL–100 ng/mL (PBS, logistic scale)；2.33 ng/mL–143.74 ng/mL (human serum, logistic scale)；检测范围: 0.1 ng/mL–100 ng/mL (PBS)；灵敏度斜率: y = 0.0025 Ln(x) + 0.0145 (PBS)；y = 0.0028 Ln(x) + 0.0010 (human serum)；R^2 = 0.9868 (PBS)；R^2 = 0.9331 (human serum)
效应效果
该传感器PBS中AFP对数浓度线性R^2=0.9868，人血清R^2=0.9331，与商品ELISA可测范围2.33–143.74 ng/mL一致。非特异BSA背景不变，非特异结合可忽略；除背景外SNR>5，未报告长期稳定性与RSD。PBS中0.1、1、10、100 ng/mL误差百分比为14.04%、16.71%、7.63%、10.50%；血清2.33 ng/mL误差44.84%，143.74 ng/mL误差6.91%。PBS检出限0.1 ng/mL（1.4 pM）优于ELISA约9.9 ng/mL，血清检测限2.33 ng/mL与ELISA相当，并避免RIA放射性风险。作者认为其成本低、可一次性使用、光学结构简单，可作临床快速免疫分析。
传感器的构成
- 基底/换能器：PMMA塑料光纤（polymethyl methacrylate, PMMA fiber，1000 μm直径），去包层区1.4 cm²，传导658 nm激光并产生倏逝波
- 表面修饰：去包层PMMA表面经化学吸附共价固定捕获抗体，形成生物识别界面
- 识别元件：鼠源单克隆抗AFP捕获抗体（anti-AFP，IgG2a，Clone C3，Sigma），特异性捕获AFP
- 封闭剂：BSA（bovine serum albumin，10 mg/mL），封闭非特异结合位点
- 信号识别探针：DyLight 649标记的鼠源单克隆抗AFP二抗（anti-AFP，IgG1，Clone F1，GeneTex），识别AFP并提供荧光
- 信号放大元件：蛋白A-金纳米颗粒（Au-PA，GNP diameter 20 nm，Sigma），通过蛋白A连接二抗，提供LSP增强并作为间隔避免荧光猝灭
- 信号读出：658 nm diode laser激发，680.4 nm窄带干涉滤光片，PMT（photomultiplier tube）检测荧光，锁相放大器提高信噪比
中文摘要
本研究报道了一种基于局域表面等离子体耦合荧光（LSPCF）的光纤生物传感器，用于直接检测人血清中的甲胎蛋白（alpha-fetoprotein, AFP）。AFP是肝细胞癌和生殖细胞肿瘤的重要肿瘤标志物，健康成人血清水平通常低于25 ng/mL，升高可提示肝癌等疾病。该传感器以去包层聚甲基丙烯酸甲酯（PMMA）光纤为基底，表面化学固定抗AFP捕获抗体，并用牛血清白蛋白（BSA）封闭；检测时采用夹心免疫分析，荧光探针由DyLight 649标记的抗AFP二抗与蛋白A-金纳米颗粒（Au-PA）组成。658 nm激光激发光纤倏逝波，进而激发金纳米颗粒表面局域表面等离子体，高效增强邻近荧光团的发射。实验结果表明，PBS中AFP在0.1–100 ng/mL范围内呈对数线性响应，人血清中2.33–143.74 ng/mL范围内亦呈线性响应；PBS检出限为0.1 ng/mL（1.4 pM），血清最低可测浓度为2.33 ng/mL，性能与ELISA相当。该传感器具有成本低、可一次性使用、光学结构简单等优点，有望用于临床快速免疫检测。
英文摘要
In this study, we demonstrated that the fiber-optic biosensor based on localized surface plasmon coupled fluorescence (LSPCF) is capable of detecting alpha-fetoprotein (AFP) in human serum. The sensitivity of LSPCF fiber-optic biosensor is not only enhanced but also the specific selectivity is improved since the fluorophores are excited by the localized surface plasmon with high efficiency. Experimentally, this fiber-optic biosensor is able to detect AFP concentration in phosphate buffered saline (PBS) solution from 0.1ng/mL to 100ng/mL whereas the linear relationship between the AFP concentrations and the fluorescence signals is shown. Furthermore, a linear response between the fluorescence signals and the concentrations of AFP in human serum from 2.33ng/mL to 143.74ng/mL is also obtained. As a result, the detection limit of the LSPCF fiber-optic biosensor on AFP detection is comparable with the conventional enzyme-linked immunosorbent assay (ELISA). Additionally, the LSPCF fiber-optic biosensor benefits on inexpensive, disposable and simpler optical geometry that can become a high efficient immunoassay comparable with the conventional ELISA and radioimmunoassay (RIA) clinically.