Development of a novel ssDNA aptamer targeting cardiac troponin I and its clinical applications
Development of a novel ssDNA aptamer targeting cardiac troponin I and its clinical applications
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DOI：
10.1007/s00216-021-03667-z
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发表时间：
2021-10-21
影响因子：
4.3
通讯作者：
Yan, Pengke
中科院分区：
文献类型：
--
作者：
Cen, Yi;Wang, Zhongping;Yan, Pengke
心肌肌钙蛋白I（cTnI）是急性心肌梗死（AMI）的特异性生物标志物。然而，用抗体制备的cTnI检测试剂盒存在许多缺陷。核酸适体是单链DNA或RNA的序列，可以克服抗体的缺陷。在此，基于适体建立了夹心ELISA方法。两种选定的ssDNA适体（Apt 3和Apt 6）显示出高结合亲和力和敏感性（Apt 3：Kd = 1.01 +/- 0.07 nM，Apt 6：k = 0.68 +/- 0.05），并且不与cTnI的相同结构域结合。因此，这两种适体可应用于DNA方法。双适体夹心法cTnI检测范围为0.05-200 ng/mL，生物分析方法验证结果符合《中国药典》（2020版）国家标准。双适体夹心ELISA法检测结果与243份血清的检测结果无显著性差异（P = 0.39，P > 0.05）。血清cTnI与ELISA联合检测的敏感性和特异性分别为96.46%和93.85%。与目前临床使用的FICA试剂盒相比，FICA法的结果更接近诊断结果。结论Apt 3和Apt 6核酸适体亲和力高、特异性强，双核酸适体夹心法检测范围广，可用于血清cTnI的检测，在AMI的诊断中具有潜在的应用价值。
Cardiac troponin I (cTnI) is a specific biomarker of acute myocardial infarction (AMI). However, cTnI detection kits prepared with antibodies have many defects. Nucleic acid aptamers are sequences of single-strand DNA or RNA that can overcome the deficiency of antibodies. Herein, sandwich ELONA methods were established based on aptamers. Two selected ssDNA aptamers (Apt3 and Apt6) showed high binding affinity and sensibility (Apt3: Kd = 1.01 +/- 0.07 nM, Apt6: k = 0.68 +/- 0.05) and did not bind to the same domain of cTnI. Therefore, these two aptamers can be applied to the ELONA methods. The detection range of cTnI using the dual-aptamer sandwich ELONA method was 0.05-200 ng/mL, and the bioanalytical method verification results can meet the national standard of Chinese Pharmacopoeia (2020 Edition). There was no difference between results of the dual-aptamer sandwich ELONA method and the diagnostic results of serum obtained from 243 people (P = 0.39, P > 0.05). The sensitivity and specificity of the ELONA with cTnI in serum were 96.46% and 93.85%, respectively. Compared with the FICA kit, which is clinically used, the consequences of ELONA method are closer to the diagnostic results. This study suggests that the aptamers Apt3 and Apt6 have high affinity and strong specificity and that the dual-aptamer sandwich ELONA method has a wide detection range and can be used to determine cTnI in serum, with potential applications in the diagnosis of AMIs.