Department of Obstetrics and Gynecology, Second Xiangya Hospital of Central South University, Changsha, 410083, China.
Mikrochim Acta. 2018 Jun 18;185(7):331. doi: 10.1007/s00604-018-2869-4.
The authors describe a method for enhancing the hybridization chain reaction (HCR) by using gold nanoparticles (AuNPs). This can considerably improve the sensitivity of electrochemical immunoassays as demonstrated for the carbohydrate antigen 125 (CA125), a biomarker for ovarian cancer. Compared to previous HCR based assays, the DNA acting as fuel strands were immobilized onto AuNPs, so that dendrimeric like chains were formed on the electrode after HCR. The improved signal is due to the reaction of DNA on the electrode. Specifically, the reaction of the phosphate groups of DNA with molybdate forms redox-active molybdophosphate, and this generates a strong electrochemical current. The immunosensor was prepared by sequential capturing, on the electrode, (a) antibody against CA125, (b) analyte (CA125), and (c) an aptamer against CA125 to form a sandwich structure. The primer on the aptamer sequence initiates HCR by annealing to one strand of DNA on the AuNPs and to another DNA in solution. The increased loading of DNA molecules onto the electrode increases the amount of phosphate groups and subsequently increases the electrical signal. The sensitivity of the assay is found to be significantly improved compared to assays without HCR and when using conventional HCR. The immunosensor was successfully applied to the determination of CA125 in human serum samples. The detection limit (based on an S/N ratio of 3) is 50 μU.mL. This indicates that this signal amplification strategy has a large potential in terms of clinical applications. It may be modified such that it also can be applied to the determination of other analytes for which proper aptamers are available. Graphical abstract Gold nanoparticle (AuNP) enhanced hybridization chain reaction is reported to improve the sensitivity of electrochemical immunosensor. Hybridization chain reaction is carried out by annealing of H1 DNA strand immobilized on AuNP to the sticky end primer sequence of the aptamer and H2 strand to the complementary sequence of H1.
作者描述了一种通过使用金纳米粒子（AuNPs）来增强杂交链式反应（HCR）的方法。这可以极大地提高电化学免疫分析的灵敏度，如针对卵巢癌标志物糖链抗原 125（CA125）的分析所证明的那样。与以前基于 HCR 的分析方法相比，作为燃料链的 DNA 被固定在 AuNPs 上，因此在 HCR 之后，树枝状链在电极上形成。改进的信号是由于电极上 DNA 的反应。具体来说，DNA 上的磷酸基团与钼酸盐反应形成具有氧化还原活性的钼磷酸，这会产生强的电化学电流。免疫传感器的制备方法是依次在电极上捕获（a）针对 CA125 的抗体，（b）分析物（CA125），和（c）针对 CA125 的适体以形成三明治结构。适体序列上的引物通过退火到 AuNPs 上的一条 DNA 链和溶液中的另一条 DNA 链来启动 HCR。DNA 分子在电极上的负载增加会增加磷酸基团的数量，从而增加电信号。与没有 HCR 的分析方法相比，该分析方法的灵敏度得到了显著提高，并且当使用常规 HCR 时也是如此。该免疫传感器成功地应用于人血清样本中 CA125 的测定。检测限（基于信噪比为 3）为 50 μU.mL。这表明这种信号放大策略在临床应用方面具有很大的潜力。可以对其进行修改，以便也可以应用于其他具有适当适体的分析物的测定。
