Departamento de Química, Universitat Politècnica de València, Camino de Vera s/n, E46022 Valencia, Spain; Instituto Interuniversitario de Investigación de Reconocimiento Molecular y Desarrollo Tecnológico (IDM), Universitat Politècnica de València-Universitat de València, Valencia, Spain; Unidad Mixta UPV-La Fe, Nanomedicine and Sensors, IIS La Fe, Valencia, Spain.
Departamento de Química, Universitat Politècnica de València, Camino de Vera s/n, E46022 Valencia, Spain.
Talanta. 2019 Jun 1;198:424-431. doi: 10.1016/j.talanta.2019.01.124. Epub 2019 Feb 10.
Consumer electronic technologies offer practical performances to develop compact biosensing systems intended for the point-of-care testing of DNA biomarkers. Herein a discrimination method for detecting single nucleotide polymorphisms, based on isothermal amplification and on-chip hybridisation, was developed and integrated into user-friendly optical devices: e.g., USB digital microscope, flatbed scanner, smartphone and DVD drive. In order to adequately identify a single base change, loop-mediated isothermal amplification (LAMP) was employed, with high yields (8 orders) within 45 min. Subsequently, products were directly hybridised to the allele-specific probes attached to plastic chips in an array format. After colorimetric staining, four consumer electronic techniques were compared. Sensitive precise measurements were taken (high signal-to-noise ratios, 10-μm image resolution, 99% scan-to-scan reproducibility). These features confirmed their potential as analytical tools, are a competitive alternative to fluorescence scanners, and incorporate additional advantages, such as user-friendly interface and connectivity for telemedicine needs. The analytical performances of the integrated platform (assay and reader) in the human samples were also excellent, with a low detection limit (100 genomic DNA copies), and reproducible (<15%) and cheap assays (< 10 €/test). The correct genotyping of a genetic biomarker (single-nucleotide polymorphism located in the GRIK4 gene) was achieved as the assigned genotypes agreed with those determined by using sequencing. The portability, favourable discriminating and read-out capabilities reveal that the implementation of mass-produced low-cost devices into minimal-specialised clinical laboratories is closer to becoming a reality.
消费类电子产品提供了实用的性能，可以开发用于即时检测 DNA 生物标志物的紧凑型生物传感系统。本文开发了一种基于等温扩增和芯片上杂交的单核苷酸多态性检测方法，并将其集成到用户友好的光学设备中：例如，USB 数字显微镜、平板扫描仪、智能手机和 DVD 驱动器。为了充分识别单个碱基变化，采用了环介导等温扩增 (LAMP)，在 45 分钟内产生 8 个数量级的高产量。随后，将产物直接杂交到以阵列形式附着在塑料芯片上的等位基因特异性探针上。经过比色染色后，比较了四种消费类电子产品。进行了灵敏、精确的测量（高信噪比、10-μm 图像分辨率、99%的扫描到扫描再现性）。这些特性证实了它们作为分析工具的潜力，是荧光扫描仪的一种有竞争力的替代品，并具有用户友好的界面和远程医疗需求的连接等额外优势。该集成平台（检测和读取器）在人类样本中的分析性能也非常出色，检测限低（100 个基因组 DNA 拷贝），重复性好（<15%）且价格便宜（<10 欧元/测试）。通过单核苷酸多态性（位于 GRIK4 基因中的单核苷酸多态性）的正确基因分型，实现了遗传生物标志物的正确基因分型，其指定的基因型与使用测序确定的基因型一致。该系统的便携性、良好的区分和读取能力表明，将大规模生产的低成本设备应用于最小化专业临床实验室更近了一步。
