With modifications to an ultra-sensitive bio-barcode (BBC) assay, we have developed a next generation aptamer-based bio-barcode (ABC) assay to detect cytochrome-c (Cyto-c), a cell death marker released from cancer cells, for anti-cancer drug screening. An aptamer is a short single-stranded DNA selected from a synthetic DNA library that is capable of binding to its target with high affinity and specificity based on its unique DNA sequence and 3D structure after folding. Similar to the BBC assay, Cyto-c is captured by a micro-magnetic particle (MMP) coated with capturing antibodies (Ab) and an aptamer specifically against Cyto-c to form sandwich structures ([MMP-Ab]–[Cyto-c]–[Aptamer]). After washing and melting, our aptamers, acting as a DNA bio-barcode, are released from the sandwiches and hybridized with the probes specially designed for RNase H for surface plasmon resonance (SPR) sensing. In an aptamer–probe duplex, RNase H digests the RNA in the probe and releases the intact aptamer for another round of hybridization and digestion. With signal enhancement effects from gold-nanorods (Au-NRs) on probes for SPR sensing, the detection limit was found to be 1 nM for the aptamer and 80 pM for Cyto-c. Without the time-consuming DNA amplification steps by PCR, the detection process of this new ABC assay can be completed within three hours. As a proof-of-concept, phenylarsine oxide was found to be a potent agent to kill liver cancer cells with multi-drug resistance at the nano-molar level. This approach thus provides a fast, sensitive and robust tool for anti-cancer drug screening.

中文翻译：

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使用SPR，RNase H和带有RNA和金纳米线的探针进行抗癌药物筛选的Aptamer生物条形码（ABC）分析

随着修改的超灵敏b IO- b AR Ç ODE（BBC）分析，我们已经制定了下一代一个基于ptamer b IO-酒吧Ç ODE（ABC）法检测cytochrome- Ç（Cyto-c），一种从癌细胞释放的细胞死亡标记，用于抗癌药物筛选。适体是选自合成DNA文库的短的单链DNA，其能够基于其独特的DNA序列和折叠后的3D结构以高亲和力和特异性结合其靶标。与BBC分析类似，Cyto-c被包裹有捕获抗体（Ab）和特异性针对Cyto-c的适体的微磁性粒子（MMP）捕获，形成夹心结构（[MMP-Ab] – [Cyto-c ] – [Aptamer]）。经过洗涤和融化后，作为DNA生物条形码的适体从三明治中释放出来，并与专为RNase H设计的探针杂交，用于表面等离振子共振（SPR）传感。在适体-探针双链体中，RNase H消化探针中的RNA，并释放完整的适体，用于另一轮杂交和消化。金纳米线（Au-NRs）对SPR感测探针具有信号增强作用，对适体的检测极限为1 nM，对Cyto-c的检测极限为80 pM。无需通过PCR进行耗时的DNA扩增步骤，这种新的ABC分析的检测过程即可在三个小时内完成。作为概念验证，发现苯ar氧化物是杀死纳摩尔浓度多药耐药性的肝癌细胞的有效剂。因此，该方法为抗癌药物筛选提供了一种快速，灵敏且强大的工具。发现适体的检出限为1 nM，Cyto-c的检出限为80 pM。无需通过PCR进行耗时的DNA扩增步骤，这种新的ABC分析的检测过程即可在三个小时内完成。作为概念验证，发现苯ar氧化物是杀死纳摩尔浓度多药耐药性的肝癌细胞的有效剂。因此，该方法为抗癌药物筛选提供了一种快速，灵敏且强大的工具。发现适体的检出限为1 nM，Cyto-c的检出限为80 pM。无需通过PCR进行耗时的DNA扩增步骤，这种新的ABC分析的检测过程即可在三个小时内完成。作为概念验证，发现苯ar氧化物是杀死纳摩尔浓度多药耐药性的肝癌细胞的有效剂。因此，该方法为抗癌药物筛选提供了一种快速，灵敏且强大的工具。