AbstractElectrochemical sandwich immunoassay strategies involving the use of carboxyl-functionalized magnetic microbeads (cMBs) and magnetic nanoparticles (cMNPs) have been evaluated and compared. The proteolytically cleaved soluble tyrosine kinase receptor sAXL was used as the target analyte. Antibodies against AXL were covalently immobilized on cMBs or cMNPs. Immunobinding of AXL was detected by means of a secondary biotinylated antibody and a streptavidin-horseradish peroxidase conjugate. The electrochemical transduction was accomplished by capturing the cMBs or cMNPs bearing the immunoconjugates onto screen-printed carbon electrodes (SPCEs) by using a small magnet. The amperometric response was measured at −0.20 V (vs the silver pseudo-reference electrode of the SPCE) upon the addition of H2O2 in the presence of hydroquinone as the redox mediator. The calibration plots for AXL extended up to 7.5 ng mL−1 when cMBs were used for the preparation of the immunosensor and up to 40 ng mL−1 in the case of using cMNPs. The respective slope values were 158 (cMBs) and 43 nA mL ng−1 (cMNPs), while the achieved LODs were 74 (cMBs) and 75 pg mL−1 (cMNPs). Although the immunosensors prepared with cMBs provided a shorter range of linearity, they exhibited a 3.7-times larger sensitivity than those constructed with cMNPs. The successful application of the new strategies was demonstrated for the determination of the endogenous content of sAXL in real human serum samples (a cut-off value of 71 ng mL−1 have been established for patients with risk of heart failure). The immunosensors constructed using cMBs or cMNPs can be advanta geously compared, in terms of sensitivity and fabrication time, with the only immunosensor for AXL previously reported. In addition, these new immunosensors took approximately half time than ELISA to perform the assay. Graphical abstractComparative evaluation of the performance of amperometric immunosensors for tyrosine kinase receptor AXL determination using carboxyl-modified magnetic microparticles (cMBs) and nanoparticles (cMNPs) and application to the determination of the endogeneous concentration in real human serum samples.

中文翻译：

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使用磁性微粒和纳米粒子的电化学免疫传感器性能的比较评估。在酪氨酸激酶受体AXL测定中的应用

摘要已经评估和比较了涉及使用羧基功能化磁微珠 (cMB) 和磁性纳米粒子 (cMNP) 的电化学夹心免疫分析策略。蛋白水解裂解的可溶性酪氨酸激酶受体 sAXL 用作目标分析物。针对 AXL 的抗体共价固定在 cMB 或 cMNP 上。通过二级生物素化抗体和链霉亲和素-辣根过氧化物酶偶联物检测 AXL 的免疫结合。电化学转导是通过使用小磁铁将带有免疫偶联物的 cMB 或 cMNP 捕获到丝网印刷碳电极 (SPCE) 上来实现的。电流响应在 -0 时测量。在氢醌作为氧化还原介质的情况下加入 H2O2 后，电压为 20 V（相对于 SPCE 的银伪参比电极）。当使用 cMB 制备免疫传感器时，AXL 的校准图扩展至 7.5 ng mL-1，在使用 cMNP 的情况下扩展至 40 ng mL-1。各自的斜率值为 158 (cMBs) 和 43 nA mL ng-1 (cMNPs)，而达到的 LOD 为 74 (cMBs) 和 75 pg mL-1 (cMNPs)。尽管用 cMBs 制备的免疫传感器提供了更短的线性范围，但它们的灵敏度比用 cMNPs 构建的传感器高 3.7 倍。新策略的成功应用被证明可用于测定真实人血清样品中 sAXL 的内源性含量（已为有心力衰竭风险的患者确定了 71 ng mL-1 的临界值）。使用 cMB 或 cMNP 构建的免疫传感器在灵敏度和制造时间方面可以与之前报道的唯一用于 AXL 的免疫传感器进行比较。此外，这些新的免疫传感器比 ELISA 花费了大约一半的时间来执行检测。图形摘要使用羧基修饰的磁性微粒 (cMB) 和纳米粒子 (cMNP) 对酪氨酸激酶受体 AXL 测定安培免疫传感器的性能进行比较评估，并应用于测定真实人血清样品中的内源性浓度。这些新的免疫传感器比 ELISA 花费了大约一半的时间来进行检测。图形摘要使用羧基修饰的磁性微粒 (cMB) 和纳米粒子 (cMNP) 对酪氨酸激酶受体 AXL 测定安培免疫传感器的性能进行比较评估，并应用于测定真实人血清样品中的内源性浓度。这些新的免疫传感器比 ELISA 花费了大约一半的时间来进行检测。图形摘要使用羧基修饰的磁性微粒 (cMB) 和纳米粒子 (cMNP) 对酪氨酸激酶受体 AXL 测定安培免疫传感器的性能进行比较评估，并应用于测定真实人血清样品中的内源性浓度。