Key Laboratory of New Molecular Diagnosis Techniques for Infectious Diseases, Institute of Radiation Medicine, 27 Taiping Road, Beijing, China.
Key Laboratory of New Molecular Diagnosis Techniques for Infectious Diseases, Institute of Radiation Medicine, 27 Taiping Road, Beijing, China.
Biosens Bioelectron. 2014 Nov 15;61:460-5. doi: 10.1016/j.bios.2014.04.018. Epub 2014 Jun 3.
We have developed a simple and sensitive assay for the detection of the RNA genetic marker associated with high pathogenicity influenza (HPAI) virus. The assay constituted of an array of Raman label tagged hairpin-DNA immobilized on a surface-enhanced Raman scattering (SERS) active substrate as the molecular sentinel (MS) reporter. Upon incubation of the assay with the target RNA, the structure of the hairpin-DNA probe changed from stem-loop configuration (closed state) to DNA/RNA hybridization configuration (open state) so that the Raman label tag will be physically separated from the SERS substrate and induce a decrease of Raman scattering intensity. A metal film over nanosphere (MFON) substrate was developed with a SERS enhancement of about 1.7 × 10(5). Based on this MS-modified substrate, the SERS signal showed a linear relationship to the target RNA in the range of 0-60 attomoles and the limit of detect is 2.67 attomoles. The non-complementary RNA sequences control was also detected and no spectral response was observed. The sensing process only required a single hybridization step and post-hybridization washing could also be omitted. Given that this ultrasensitive biosensor assay is free of polymerase chain reaction (PCR) amplification, it would be a potential diagnostic tool for point-of-care HPAI virus detection.
我们开发了一种简单而灵敏的检测方法，用于检测与高致病性流感（HPAI）病毒相关的 RNA 遗传标志物。该检测方法由一系列固定在表面增强拉曼散射（SERS）活性衬底上的 Raman 标记发夹 DNA 组成，作为分子探针（MS）报告器。将该检测方法与靶 RNA 孵育后，发夹 DNA 探针的结构从茎环构象（闭合状态）转变为 DNA/RNA 杂交构象（开放状态），从而使 Raman 标记标签将与 SERS 衬底物理分离，并诱导拉曼散射强度降低。我们开发了一种具有约 1.7×10(5) SERS 增强的金属膜覆盖纳米球（MFON）衬底。基于这种 MS 修饰的衬底，SERS 信号与靶 RNA 在 0-60 飞摩尔范围内呈线性关系，检测限为 2.67 飞摩尔。非互补 RNA 序列对照也被检测到，没有观察到光谱响应。该传感过程仅需要单个杂交步骤，杂交后也可以省略洗涤步骤。鉴于这种超灵敏的生物传感器检测方法无需聚合酶链反应（PCR）扩增，它将成为一种用于即时护理 HPAI 病毒检测的潜在诊断工具。
