Yu Shi-Song, Shi Yu-Jun, Wang Di, Qiang Ti-Ti, Zhao Ya-Qi, Wang Xin-Yu, Zhao Jia-Meng, Dong Lin-Yi, Huang Ya-Jie, Wang Xian-Hua
Tianjin Key Laboratory on Technologies Enabling Development of Clinical Therapeutics and Diagnostics, School of Pharmacy, Tianjin Medical University, Tianjin, 300070, China.
Tianjin Key Laboratory on Technologies Enabling Development of Clinical Therapeutics and Diagnostics, School of Pharmacy, Tianjin Medical University, Tianjin, 300070, China.
Anal Chim Acta. 2023 Jun 8;1259:341202. doi: 10.1016/j.aca.2023.341202. Epub 2023 Apr 8.
Determination of trace glycoprotein has important guiding significance in clinical diagnosis and is usually achieved by immunoaffinity. However, immunoaffinity possesses inherent drawbacks, such as poor probability of high-quality antibodies, instability of biological reagents, and harmfulness of chemical labels to the body. Herein, we propose an innovative method of peptide-oriented surface imprinting to fabricate artificial antibody for recognition of glycoprotein. By integrating peptide-oriented surface imprinting and PEGylation, an innovative hydrophilic peptide-oriented surface imprinting magnetic nanoparticle (HPIMN) was successfully fabricated with human epidermal growth factor receptor-2 (HER2) as a model glycoprotein template. In addition, we further prepared a novel boronic acid-modified/fluorescein isothiocyanate-loaded/polyethylene glycol-covered carbon nanotube (BFPCN) as fluorescence signal output device, which was loaded with numerous fluorescent molecules could specifically label the cis-diol of glycoprotein at physiological pH via boronate-affinity interaction. To prove the practicability, we proposed a HPIMN-BFPCN strategy, in which the HPIMN first selectively captured the HER2 due to the molecular imprinted recognition and then the BFPCN specific labeled the exposed cis-diol of HER2 based on the boronate-affinity reaction. The HPIMN-BFPCN strategy exhibited ultrahigh sensitivity with limit of detection of 14 fg mL and was successfully used in the determination of HER2 in spiked sample with recovery and relative standard deviation in the range of 99.0%-103.0% and 3.1%-5.6%, respectively. Therefore, we believe that the novel peptide-oriented surface imprinting has great potential to become an universal strategy for fabrication of recognition units for other protein biomarkers, and the synergy sandwich assay could become a powerful tool in prognosis evaluation and clinical diagnosis of glycoprotein-related diseases.
痕量糖蛋白的测定在临床诊断中具有重要的指导意义，通常通过免疫亲和法实现。然而，免疫亲和法存在固有缺陷，如高质量抗体的获得概率低、生物试剂不稳定以及化学标记对人体的危害性。在此，我们提出一种创新的肽导向表面印迹方法来制备用于识别糖蛋白的人工抗体。通过整合肽导向表面印迹和聚乙二醇化，以人表皮生长因子受体2（HER2）作为模型糖蛋白模板，成功制备了一种创新的亲水性肽导向表面印迹磁性纳米颗粒（HPIMN）。此外，我们进一步制备了一种新型的硼酸修饰/异硫氰酸荧光素负载/聚乙二醇包覆的碳纳米管（BFPCN）作为荧光信号输出装置，其负载有大量荧光分子，可在生理pH条件下通过硼酸亲和相互作用特异性标记糖蛋白的顺式二醇。为证明其实用性，我们提出了一种HPIMN-BFPCN策略，其中HPIMN首先由于分子印迹识别选择性捕获HER2，然后BFPCN基于硼酸亲和反应特异性标记HER2暴露的顺式二醇。HPIMN-BFPCN策略表现出超高灵敏度，检测限为14 fg/mL，并成功用于加标样品中HER2的测定，回收率和相对标准偏差分别在99.0%-103.0%和3.1%-5.6%范围内。因此，我们认为这种新型的肽导向表面印迹有很大潜力成为制备其他蛋白质生物标志物识别单元的通用策略，并且协同夹心分析可能成为糖蛋白相关疾病预后评估和临床诊断的有力工具。
