College of Life Sciences, Northwest University, Xi'an, 710069, China.
Nanoscale. 2020 May 14;12(18):10098-10105. doi: 10.1039/d0nr00360c.
Rapid and simple detection of single nucleotide polymorphism (SNP) is vital for individualized diagnosis and eventual treatment in the current clinical setting. In this study, we developed a tetra-primer ARMS-PCR combined lateral flow assay (T-ARMS-PCR-LFA) method for simultaneous visual detection of two alleles. By using four primers labeled with digoxin, biotin and Cy5 separately in one PCR reaction, the amplified allele-specific products could be captured by streptavidin and the anti-Cy5 antibody on two separated test lines of a LFA strip, which allows the presentation of both alleles within the single LFA strip. Both DNA and whole blood can be used as templates in this genotyping method in which the whole detection process is completed within 75 minutes. The performance assay of T-ARMS-PCR-LFA demonstrates the accuracy, specificity and sensitivity of this method. One hundred human whole blood samples were used for MTHFR C677T genotyping in T-ARMS-PCR-LFA. The concordance rate of the results detected was up to 100% when compared with that of the sequencing results. Collectively, this newly developed method is highly applicable for SNP screening in clinical practices.
快速、简便地检测单核苷酸多态性（SNP）对于当前临床环境中的个体化诊断和最终治疗至关重要。在本研究中，我们开发了一种四引物 ARMS-PCR 结合侧流分析（T-ARMS-PCR-LFA）方法，用于同时可视化检测两个等位基因。通过在一个 PCR 反应中分别使用四个标记有地高辛、生物素和 Cy5 的引物，扩增的等位基因特异性产物可以被链霉亲和素和 LFA 条上两个分离的测试线上的抗 Cy5 抗体捕获，从而在单个 LFA 条上呈现两个等位基因。该基因分型方法既可以使用 DNA 作为模板，也可以使用全血作为模板，整个检测过程在 75 分钟内完成。T-ARMS-PCR-LFA 的性能评估表明了该方法的准确性、特异性和灵敏度。100 个人全血样本用于 T-ARMS-PCR-LFA 中的 MTHFR C677T 基因分型。与测序结果相比，检测结果的符合率高达 100%。总的来说，这种新开发的方法非常适用于临床实践中的 SNP 筛查。
