组成图示
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传感器类型
荧光生物传感器
检测对象
葡萄糖（glucose，β-D-glucose）；样品基质：去离子水（DI water）、模拟组织间液（simulated interstitial fluid, SIF）
检测原理
该传感器基于脱辅基葡萄糖氧化酶（apo-GOx）的亲和结合而非酶催化。apo-GOx去除FAD后不再氧化葡萄糖，但保留对β-D-葡萄糖的结合特异性，从而避免底物消耗和过氧化氢等副产物。FITC-右旋糖酐与TRITC-apo-GOx结合时，FITC作为供体向TRITC受体发生FRET，使FITC发射降低、TRITC发射增强。当葡萄糖进入溶解核海藻酸微球后，与apo-GOx竞争结合并置换FITC-右旋糖酐，FRET效率下降，FITC/TRITC发射峰比升高。该峰比随葡萄糖浓度增加而增大，在0–30 mM范围内响应，0–6 mM近似线性；比例荧光读数可校正光源波动和传感器浓度变化。
检测灵敏度
灵敏度: 0.52%/mM glucose in DI water；0.6%/mM glucose in simulated interstitial fluid；分析响应范围: 0–30mM glucose；线性范围: 0–6mM glucose（linear up to 6mM glucose concentration）
效应效果
选择性方面，apo-GOx对β-D-葡萄糖结合特异，低浓度甘露糖、果糖、蔗糖和α-D-葡萄糖不显著干扰。可逆性方面，葡萄糖去除后FITC/TRITC峰比恢复基线，平均基线漂移在DI水中为1.03%（SD 0.61），在SIF中为1.3%（SD 0.67）。动态响应时间最长约2 min。重复性SD为1%，三批传感器重现性SD为1.5%。SIF中连续测试30天，灵敏度保持0.6%/mM，SD为1.1%；[PAH/PSS]2涂层15 h泄漏约4%，优于[PLL/HA]2的9%和[DEAE/CS]2的7%。L929细胞毒性实验显示存活率约100%。作者主张用于植入式连续葡萄糖监测。
传感器的构成
- 微球基底：低粘度海藻酸钠（alginate）经CaCl2离子交联形成海藻酸微球，作为封装传感试剂的载体，并经柠檬酸钠部分去交联形成溶解核。
- 识别元件：TRITC标记的葡萄糖氧化酶脱辅基蛋白（TRITC-apo-GOx）作为葡萄糖结合蛋白，保留β-D-葡萄糖结合特异性但不催化氧化。
- 信号标记物：FITC标记右旋糖酐（FITC-dextran，FD，优选500 kDa）作为FRET供体，与TRITC-apo-GOx结合形成竞争结合检测体系。
- 纳米薄膜修饰层：层层自组装[PAH/PSS]2聚电解质多层纳米薄膜（聚烯丙基胺盐酸盐PAH、聚苯乙烯磺酸钠PSS），稳定去交联微球并限制传感试剂泄漏。
- 信号读出：荧光分光光度计（Hitachi F-2500）配合石英微流动池和注射泵，连续记录FITC与TRITC发射峰比值。
中文摘要
微粒光学传感器有望作为植入式智能材料用于体内分析。本文评估了含有均相荧光共振能量转移（FRET）竞争结合葡萄糖检测体系的溶解核海藻酸微球的可逆响应。采用层层自组装技术在含检测体系的海藻酸微球表面沉积多层纳米薄膜涂层，使海藻酸去交联后传感器体系保持稳定。随后在去离子水和模拟组织间液中，利用流动池建立受控动态流动条件，测定微球对葡萄糖的响应。动态条件下，葡萄糖灵敏度在去离子水中为0.52%/mM，在模拟组织间液中为0.6%/mM；两种体系的分析响应范围均为0–30 mM葡萄糖，覆盖正常血糖及高血糖、低血糖等病理范围。该传感器在动态流动条件下连续测试一个月，表现出可重复且可重现的响应。L929小鼠成纤维细胞体外细胞毒性实验表明，具有纳米薄膜涂层的溶解核海藻酸微球传感器系统具有足够生物相容性，可作为植入式葡萄糖生物传感器使用。
英文摘要
Microparticle optical sensors hold potential as implantable smart materials for in vivo analysis. In this work, the reversible response of dissolved-core alginate microspheres containing a homogeneous fluorescence resonance energy transfer (FRET)-based competitive binding assay for glucose was evaluated. The layer-by-layer self assembly technique was used to deposit multilayered nanofilm coatings on the alginate microspheres containing the assay, thereby stabilizing the sensor system when the alginate was de-crosslinked. The response to glucose was then determined in DI water and simulated interstitial fluid (SIF) using a flow cell to establish controlled, dynamic flow conditions for demonstrating reversibility. The glucose sensitivity under dynamic conditions was estimated to be 0.52%/mM glucose in DI water and 0.6%/mM glucose in simulated interstitial fluid; in both cases, the analytical response range was 0-30 mM glucose, covering both physiological (normoglycemia) and pathophysiological range (hyperglycemia and hypoglycemia). The sensor demonstrated a repeatable and reproducible response when tested over a period of one month, under dynamic flow conditions. Finally, in vitro cytotoxicity assays performed with L929 mouse fibroblast cell lines suggested that the dissolved-core alginate microsphere sensor system with nanofilm coating has sufficient biocompatibility for use as implantable glucose biosensors.