State Key Laboratory of Bioelectronics, School of Biological Science and Medical Engineering, Southeast University, Nanjing 210096, China.
Biosens Bioelectron. 2010 Dec 15;26(4):1442-8. doi: 10.1016/j.bios.2010.07.078. Epub 2010 Jul 30.
A sensitive and selective biosensor platform suited for SNP type using Fe@Au magnetic nanoparticles (GMNPs) to fabricate bead array is described. This new platform integrates the rapid binding kinetics of magnetic nanoparticles carriers, the multiplexing and encoding capabilities of chips, and tagged array. As a DNA sensor, the biotinylated single-stranded DNA was obtained by asymmetry PCR amplification, and then captured by GMNPs modified with streptavidin to form GMNP-ssDNA complexes without further purification. The complexes were immobilized on the slide to fabricate bead array through magnetic field. The bead array was hybridized with the corresponding allele-specific tag probes for each locus, and a pair of given universal detectors were applied to these markers analysis. Using bead array, all samples can be analyzed in one hybridization chamber which lowers the cost of the assay. Using universal tags, only a pair of universal dual-color probes labeled fluorophores was used for multiplex genotyping. Without the need of laborious and time-consuming elution, the experiment process was simple, reproducible and easy to handle. Two SNPs loci from 12 individual samples were discriminated using this platform and the results demonstrated that the expected scores and good discrimination were obtained between the two alleles from the two SNP loci. In summary, the integrated sensitive platform is adaptable and versatile, while offering a high-throughput capability needed for genome research and clinical applications.
本文描述了一种适用于 SNP 类型的灵敏和选择性生物传感器平台,该平台使用 Fe@Au 磁性纳米颗粒 (GMNPs) 来制备珠阵列。这个新平台集成了磁性纳米颗粒载体的快速结合动力学、芯片的多重化和编码能力以及标记的阵列。作为 DNA 传感器,通过不对称 PCR 扩增获得生物素化的单链 DNA,然后通过链霉亲和素修饰的 GMNPs 捕获,形成 GMNP-ssDNA 复合物,无需进一步纯化。通过磁场将复合物固定在载玻片上,以形成珠阵列。珠阵列与每个位点的相应等位基因特异性标记探针杂交,并应用一对给定的通用探测器进行这些标记物的分析。使用珠阵列,可以在一个杂交室内分析所有样本,从而降低了检测成本。使用通用标签,仅需一对标记有荧光染料的通用双色探针即可进行多重基因分型。无需费力和耗时的洗脱,实验过程简单、可重复且易于操作。使用该平台区分了来自 12 个个体样本的两个 SNP 位点,结果表明,从两个 SNP 位点的两个等位基因获得了预期的分数和良好的区分。总之,集成的灵敏平台适应性强,功能多样,同时提供了基因组研究和临床应用所需的高通量能力。