Department of Chemistry, Faculty of Science, Srinakharinwirot University , Bangkok, 10110, Thailand.
Nanoelectronics and MEMS Laboratory, National Electronics and Computer Technology Center , Pathumthani 12120, Thailand.
Anal Chem. 2017 May 16;89(10):5428-5435. doi: 10.1021/acs.analchem.7b00255. Epub 2017 Apr 27.
The development of simple fluorescent and colorimetric assays that enable point-of-care DNA and RNA detection has been a topic of significant research because of the utility of such assays in resource limited settings. The most common motifs utilize hybridization to a complementary detection strand coupled with a sensitive reporter molecule. Here, a paper-based colorimetric assay for DNA detection based on pyrrolidinyl peptide nucleic acid (acpcPNA)-induced nanoparticle aggregation is reported as an alternative to traditional colorimetric approaches. PNA probes are an attractive alternative to DNA and RNA probes because they are chemically and biologically stable, easily synthesized, and hybridize efficiently with the complementary DNA strands. The acpcPNA probe contains a single positive charge from the lysine at C-terminus and causes aggregation of citrate anion-stabilized silver nanoparticles (AgNPs) in the absence of complementary DNA. In the presence of target DNA, formation of the anionic DNA-acpcPNA duplex results in dispersion of the AgNPs as a result of electrostatic repulsion, giving rise to a detectable color change. Factors affecting the sensitivity and selectivity of this assay were investigated, including ionic strength, AgNP concentration, PNA concentration, and DNA strand mismatches. The method was used for screening of synthetic Middle East respiratory syndrome coronavirus (MERS-CoV), Mycobacterium tuberculosis (MTB), and human papillomavirus (HPV) DNA based on a colorimetric paper-based analytical device developed using the aforementioned principle. The oligonucleotide targets were detected by measuring the color change of AgNPs, giving detection limits of 1.53 (MERS-CoV), 1.27 (MTB), and 1.03 nM (HPV). The acpcPNA probe exhibited high selectivity for the complementary oligonucleotides over single-base-mismatch, two-base-mismatch, and noncomplementary DNA targets. The proposed paper-based colorimetric DNA sensor has potential to be an alternative approach for simple, rapid, sensitive, and selective DNA detection.
基于吡咯烷二肽核酸（acpcPNA）诱导纳米颗粒聚集的纸基比色法用于 DNA 检测：一种替代传统比色法的方案
由于此类检测在资源有限的环境中具有实用性，因此能够实现即时检测（point-of-care，POC）的简单荧光和比色测定法的开发一直是一个重要的研究课题。最常见的模式是利用与互补检测链的杂交，再加上灵敏的报告分子。本文报道了一种基于吡咯烷二肽核酸（acpcPNA）诱导纳米颗粒聚集的纸基比色法用于 DNA 检测，作为传统比色法的替代方案。与 DNA 和 RNA 探针相比，肽核酸（PNA）探针是一种很有吸引力的替代品，因为它们具有化学和生物稳定性，易于合成，并且与互补的 DNA 链高效杂交。acpcPNA 探针在 C 末端带有一个来自赖氨酸的正电荷，在没有互补 DNA 的情况下会引起柠檬酸根稳定的银纳米颗粒（AgNPs）的聚集。在存在靶 DNA 的情况下，由于静电排斥，形成带负电荷的 DNA-acpcPNA 双链体导致 AgNPs 分散，从而产生可检测的颜色变化。本文研究了影响该检测方法灵敏度和选择性的因素，包括离子强度、AgNP 浓度、PNA 浓度和 DNA 链错配。根据上述原理，使用该方法筛选了合成的中东呼吸综合征冠状病毒（MERS-CoV）、结核分枝杆菌（MTB）和人乳头瘤病毒（HPV）DNA，开发了基于纸的比色分析装置。通过测量 AgNPs 的颜色变化来检测寡核苷酸靶标，得出的检测限分别为 1.53（MERS-CoV）、1.27（MTB）和 1.03 nM（HPV）。acpcPNA 探针对互补寡核苷酸表现出高选择性，而对单碱基错配、双碱基错配和非互补 DNA 靶标则没有选择性。该基于纸的比色 DNA 传感器具有作为简单、快速、灵敏和选择性 DNA 检测替代方法的潜力。
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