Nanobioelectronics & Biosensors Group, CIN2 (ICN-CSIC), Catalan Institute of Nanotechnology, Campus UAB, Bellaterra, (Barcelona), Spain.
Biosens Bioelectron. 2013 Feb 15;40(1):24-31. doi: 10.1016/j.bios.2012.05.021. Epub 2012 May 31.
A high sensitive voltammetric method for rapid determination of thrombin spiked in whole blood by taking advantage of both aptamer-based recognition and the use of a nanoporous membrane has been developed. The nanoporous membrane not only acts as platform for the thrombin recognition but also as filter of the micrometric components such as white and red blood cells, consequently minimizing matrix effects. The protocol involves a sandwich format in the inner walls (200 nm diameter) of an anodized alumina oxide filter membrane (AAO). The analytical signal, by DPV oxidation of Fe(CN)(6), is based on the blockage in the pores which affects the diffusion of Fe(CN)(6) to the screen-printed carbon electrotransducer (SPCEs) modified with the membrane. By labeling the anti-thrombin IgG with AuNPs followed by silver enhancement a greater passive signal enhancement in comparison to the membrane blockage has been observed. The contribution of both electrostatic/steric effects in this blockage due to the subsequent formation of the aptamer-thrombin complex and the final sandwich assay is investigated. The efficiency of the system is also monitored by microscopic techniques. The resulted biosensing system allows detecting thrombin spiked in whole blood at very low levels (LOD 1.8 ng mL(-1)) which are within the range of clinical interest for the diagnostic of coagulation abnormalities as well as pulmonary metastasis.
利用基于适配体的识别和使用纳米多孔膜的优势，开发了一种高灵敏度的伏安法，可快速测定全血中添加的凝血酶。纳米多孔膜不仅作为凝血酶识别的平台，而且作为微米级成分（如白细胞和红细胞）的过滤器，从而最大限度地减少基质效应。该方案涉及在氧化铝氧化膜（AAO）过滤器的内壁（200nm 直径）中形成三明治结构。分析信号是通过Fe(CN)(6)的 DPV 氧化产生的，这基于阻塞在影响Fe(CN)(6)扩散到修饰有膜的丝网印刷碳电极（SPCE）的孔中。通过将抗凝血酶 IgG 与 AuNPs 标记，然后进行银增强，与膜阻塞相比，观察到更大的被动信号增强。还研究了由于适配体-凝血酶复合物的后续形成以及最终三明治测定而导致的这种阻塞中静电/空间效应的贡献。该系统的效率也通过微观技术进行监测。所得到的生物传感系统可检测全血中极低水平（LOD 1.8ng mL(-1))的凝血酶，这在诊断凝血异常以及肺转移方面具有临床意义。
