Jo Hunho, Gu Hyunwoo, Jeon Weejeong, Youn Hyungjun, Her Jin, Kim Seong-Kyeong, Lee Jeongbong, Shin Jae Ho, Ban Changill
Department of Chemistry, Pohang University of Science and Technology , 77, Cheongam-Ro, Nam-Gu, Pohang, Gyeongbuk 790-784, South Korea.
Department of Chemistry, Kwangwoon University , Wolgye-Dong, Nowon-Gu, Seoul 139-701, South Korea.
Anal Chem. 2015 Oct 6;87(19):9869-75. doi: 10.1021/acs.analchem.5b02312. Epub 2015 Sep 30.
Cardiac troponin I (cTnI) is well-known as a promising biomarker for the early diagnosis of acute myocardial infarction (AMI). In this work, single-stranded DNA aptamers against cTnI were identified by the Systematic Evolution of Ligands by Exponential enrichment (SELEX) method. The aptamer candidates exhibited a high selectivity and sensitivity toward both cTnI and the cardiac Troponin complex. The binding affinities of each aptamer were evaluated based on their dissociation constants (Kd) by surface plasma resonance. The Tro4 aptamer that had the highest binding capacity to cTnI showed a very low Kd value (270 pM) compared with that of a cTnI antibody (20.8 nM). Furthermore, we designed a new electrochemical aptasensor based on square wave voltammetry using ferrocene-modified silica nanoparticles. The developed aptasensor demonstrated an excellent analytical performance for cTnI with a wide linear range of 1-10 000 pM in a buffer and a detection limit of 1.0 pM (24 pg/mL; S/N = 3), which was noticeably lower than the cutoff values (70-400 pg/mL). The specificity of the aptamers was also examined using nontarget proteins, demonstrating that the proposed sensor responded to only cTnI. In addition, cTnI was successfully detected in a human serum albumin solution. On the basis of the calibration curve that was constructed, the concentrations of cTnI in a solution supplemented with human serum were effectively measured. The calculated values correlated well with the actual concentrations of cTnI. It is anticipated that the highly sensitive and selective aptasensor for cTnI could be readily applicable for the accurate diagnosis of AMI.
心肌肌钙蛋白I（cTnI）作为急性心肌梗死（AMI）早期诊断的一种很有前景的生物标志物而广为人知。在这项工作中，通过指数富集配体系统进化（SELEX）方法鉴定了针对cTnI的单链DNA适配体。候选适配体对cTnI和心肌肌钙蛋白复合物均表现出高选择性和敏感性。基于表面等离子体共振的解离常数（Kd）评估了每个适配体的结合亲和力。与cTnI抗体（20.8 nM）相比，对cTnI具有最高结合能力的Tro4适配体显示出非常低的Kd值（270 pM）。此外，我们设计了一种基于方波伏安法的新型电化学适配体传感器，该传感器使用二茂铁修饰的二氧化硅纳米颗粒。所开发的适配体传感器对cTnI表现出优异的分析性能，在缓冲液中的线性范围为1 - 10000 pM，检测限为1.0 pM（24 pg/mL；S/N = 3），明显低于临界值（70 - 400 pg/mL）。还使用非靶蛋白检查了适配体的特异性，结果表明所提出的传感器仅对cTnI有响应。此外，在人血清白蛋白溶液中成功检测到了cTnI。根据构建的校准曲线，有效地测量了添加人血清的溶液中cTnI的浓度。计算值与cTnI的实际浓度相关性良好。预计用于cTnI的高灵敏度和高选择性适配体传感器可很容易地应用于AMI的准确诊断。
