A direct detection of Escherichia coli genomic DNA using gold nanoprobes.
A direct detection of Escherichia coli genomic DNA using gold nanoprobes.
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使用金纳米探针直接检测大肠杆菌基因组 DNA。
DOI：
10.1186/1477-3155-10-8
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发表时间：
2012-02-06
影响因子：
10.2
通讯作者：
Jaffar Ali BM
中科院分区：
文献类型：
--
作者：
Padmavathy B;Vinoth Kumar R;Jaffar Ali BM
在临床和法医样品的诊断等情况下，需要高度灵敏、快速和特异的DNA检测方法。虽然使用PCR的常规DNA扩增可以提供快速结果，但它在诊断实验室中没有广泛应用，部分原因是它需要熟练的人员和昂贵的设备。为了克服这些限制，纳米颗粒已被探索作为信号探针，用于超灵敏的DNA检测，可用于现场应用。在纳米材料中，金纳米颗粒（AuNPs）已被广泛使用，主要是因为其光学性质和与各种生物分子功能化的能力。我们报告了一种使用单链寡核苷酸功能化的金纳米颗粒（AuNP-oligo探针）作为视觉检测探针用于快速和特异性检测大肠杆菌的方案。与含有互补序列的靶DNA杂交的AuNP-oligo探针保持红色，而没有与探针互补的DNA序列的测试样品由于酸诱导的AuNP-oligo探针聚集而变成紫色.通过肉眼观察溶液的颜色变化，证明从其基因组DNA直接和快速检测致病性大肠杆菌，而不需要PCR扩增。未扩增基因组DNA的检测限为~54 ng。该方法在基因组DNA提取后需要不到30分钟即可完成。然而，通过使用未扩增的酶消化的基因组DNA，达到11.4 ng的检测限。紫外-可见光谱测量和原子力显微镜成像的结果进一步支持基于聚集的视觉辨别的假设。为了阐明其在医学诊断中的实用性，该检测方法在从当地医院获得的致病性大肠杆菌临床菌株和加标尿样上进行了验证。结果表明，该方法敏感性为100%，特异性高，与非大肠杆菌菌株无交叉反应。这项工作进入了一类新的DNA/金纳米颗粒混合材料，该材料可能具有可以控制的光学性质，可用于诊断应用。我们注意到，应该可以很容易地扩展这种策略，以开发用于即时检测的新型DNA生物传感器。该方法的显著特点包括低成本、稳健的试剂和简单的病原体比色检测。
In situation like diagnosis of clinical and forensic samples there exists a need for highly sensitive, rapid and specific DNA detection methods. Though conventional DNA amplification using PCR can provide fast results, it is not widely practised in diagnostic laboratories partially because it requires skilled personnel and expensive equipment. To overcome these limitations nanoparticles have been explored as signalling probes for ultrasensitive DNA detection that can be used in field applications. Among the nanomaterials, gold nanoparticles (AuNPs) have been extensively used mainly because of its optical property and ability to get functionalized with a variety of biomolecules. We report a protocol for the use of gold nanoparticles functionalized with single stranded oligonucleotide (AuNP- oligo probe) as visual detection probes for rapid and specific detection of Escherichia coli. The AuNP- oligo probe on hybridization with target DNA containing complementary sequences remains red whereas test samples without complementary DNA sequences to the probe turns purple due to acid induced aggregation of AuNP- oligo probes. The color change of the solution is observed visually by naked eye demonstrating direct and rapid detection of the pathogenic Escherichia coli from its genomic DNA without the need for PCR amplification. The limit of detection was ~54 ng for unamplified genomic DNA. The method requires less than 30 minutes to complete after genomic DNA extraction. However, by using unamplified enzymatic digested genomic DNA, the detection limit of 11.4 ng was attained. Results of UV-Vis spectroscopic measurement and AFM imaging further support the hypothesis of aggregation based visual discrimination. To elucidate its utility in medical diagnostic, the assay was validated on clinical strains of pathogenic Escherichia coli obtained from local hospitals and spiked urine samples. It was found to be 100% sensitive and proves to be highly specific without any cross reaction with non-Escherichia coli strains. This work gives entry into a new class of DNA/gold nanoparticles hybrid materials which might have optical property that can be controlled for application in diagnostics. We note that it should be possible to extend this strategy easily for developing new types of DNA biosensor for point of care detection. The salient feature of this approach includes low-cost, robust reagents and simple colorimetric detection of pathogen.
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影响因子：
2.2
作者：
Liandris, Emmanouil;Gazouli, Maria;Ikonomopoulos, John
通讯作者：
Ikonomopoulos, John
使用金纳米探针进行 RNA 定量——在癌症诊断中的应用。
影响因子：
10.2
作者：
Conde J;de la Fuente JM;Baptista PV
通讯作者：
Baptista PV
影响因子：
15
作者：
Cao, YW;Jin, R;Mirkin, CA
通讯作者：
Mirkin, CA
影响因子：
7.4
作者：
Lee, Jae-Seung;Mirkin, Chad A.
通讯作者：
Mirkin, Chad A.
影响因子：
64.8
作者：
Mirkin, CA;Letsinger, RL;Storhoff, JJ
通讯作者：
Storhoff, JJ