组成图示
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传感器类型
综述或非传感器论文
检测对象
HER2（人表皮生长因子受体2，HER2/ErbB2）；样品基质：HER2阳性SKOV-3细胞、LS174T/SKOV-3异种移植瘤、小鼠组织/血液、小鼠血清。
检测原理
本文并非传统生物传感器，而是放射性分子影像探针。合成Affibody分子ZHER2:342通过三螺旋结构域特异性识别HER2胞外域；MAG3序列在N端位点特异性螯合99mTc，形成99mTc-MAG3-ZHER2:342。探针与肿瘤细胞表面HER2结合后，放射性核素99mTc衰变发射γ光子，被伽马相机探测；HER2表达越高，结合探针越多，肿瘤局部γ计数越高，从而形成肿瘤-背景对比。SPR表征中，HER2或HSA固定于CM5芯片，Affibody流过时结合引起界面折射率变化，Biacore 2000以响应单位RU读出结合动力学。该体系无酶催化或核酸放大，信号放大主要依赖放射性标记与高亲和力识别。
检测灵敏度
原文未报告 LOD、线性范围、灵敏度斜率或相关系数。
效应效果
MAG3-ZHER2:342纯度>90%，SPR Kd 200 pM。99mTc标记效率82±12%，胶体<2%，纯化后放射性纯度>98%；300倍半胱氨酸1 h后87.5±1.8%仍结合，血清1 h后>90%完整。SKOV-3细胞结合可被未标记ZHER2:342阻断，从50.4±0.2%降至1.5±0.02%，ABC 86±1%；24 h细胞滞留55%，高于125I-PIB约40%。LS174T瘤2–6 h摄取约7%IA/g，阻断组4 h由6.12±2.02降至0.28±0.05%IA/g（p=0.001），瘤/血比2、4、6 h为12.5、23.8、39.3。与125I-PIB相比肿瘤摄取相近；6 h显影清晰，24 h肿瘤/肾显影。作者认为可用于HER2肿瘤显像及治疗选择。
传感器的构成
- 非传感器基底：Biacore 2000 SPR 仪器与 CM5 传感器芯片，仅用于结合表征
- 非传感器固定层：CM5 芯片表面，固定重组 HER2 胞外域或 HSA
- 非传感器识别元件：重组 HER2 胞外域（HER2）或 HSA，用于捕获 Affibody 分子
- 非传感器分析物：ZHER2:342 或 MAG3-ZHER2:342 Affibody 分子，作为结合探针
- 非传感器信号标记：99mTc-MAG3-ZHER2:342 放射性标记物，用于伽马相机成像
- 非传感器读出：SPR 响应单位（RU）或伽马相机 γ 计数，反映结合量或放射性分布
中文摘要
人表皮生长因子受体2（HER2）在恶性肿瘤中的表达具有明确的预后和预测价值，非侵入性显像可为患者管理提供重要诊断信息。本研究旨在开发一种利用含锝-99m螯合序列巯基乙酰-甘氨酰-甘氨酰-甘氨酰（MAG3）标记抗HER2 Affibody分子ZHER2:342的方法，并评价其靶向性能。作者采用Fmoc/tBu固相肽合成组装MAG3-ZHER2:342，通过反相高效液相色谱、电喷雾质谱、生物传感器分析和圆二色光谱进行生化表征，并建立含钠/钾酒石酸的99mTc标记流程。结果显示，合成MAG3-ZHER2:342对HER2受体的亲和力为0.2 nM，99mTc标记效率约75–80%；标记物在体外仍特异性结合HER2阳性SKOV-3细胞。在携带LS174T异种移植瘤的小鼠中，99mTc-MAG3-ZHER2:342表现出与放射性碘类似物相近的特异性肿瘤靶向和对比度，伽马相机显像可清晰、特异性地显示HER2表达。结论表明，在化学合成中引入含巯基乙酰的螯合序列可实现ZHER2:342的位点特异性99mTc标记，并保持靶向能力。
英文摘要
PURPOSE: Expression of human epidermal growth factor receptor type 2 (HER2) in malignant tumours possesses well-documented prognostic and predictive value. Non-invasive imaging of expression can provide valuable diagnostic information, thereby influencing patient management. Previously, we reported a phage display selection of a small (about 7 kDa) protein, the Affibody molecule Z(HER2:342), which binds HER2 with subnanomolar affinity, and demonstrated the feasibility of targeting of HER2-expressing xenografts using radioiodinated Z(HER2:342). The goal of this study was to develop a method for (99m)Tc labelling of Z(HER2:342) using the MAG3 chelator, which was incorporated into Z(HER2:342) using peptide synthesis, and evaluate the targeting properties of the labelled conjugate.
METHODS: MAG3-Z(HER2:342) was assembled using Fmoc/tBu solid phase peptide synthesis. Biochemical characterisation of the agent was performed using RP-HPLC, ESI-MS, biosensor studies and circular dichroism. A procedure for (99m)Tc labelling in the presence of sodium/potassium tartrate was established. Tumour targeting was evaluated by biodistribution study and gamma camera imaging in xenograft-bearing mice. Biodistribution of (99m)Tc-MAG3-Z(HER2:342) and (125)I-para-iodobenzoate -Z(HER2:342) was compared 6 h p.i.
RESULTS: Synthetic MAG3-Z(HER2:342) possessed an affinity of 0.2 nM for HER2 receptors. The peptide was labelled with (99m)Tc with an efficiency of about 75-80%. Labelled (99m)Tc-MAG3-Z(HER2:342) retained capacity to bind specifically HER2-expressing SKOV-3 cells in vitro. (99m)Tc-MAG3-Z(HER2:342) showed specific tumour targeting with a contrast similar to a radioiodinated analogue in mice bearing LS174T xenografts. Gamma camera imaging demonstrated clear and specific visualisation of HER2 expression.
CONCLUSION: Incorporation of a mercaptoacetyl-containing chelating sequence during chemical synthesis enabled site-specific (99m)Tc labelling of the Z(HER2:342) Affibody molecule with preserved targeting capacity.