Direct Detection of Low Abundance Genes of Single Point Mutation

• DOI: 10.1021/acs.nanolett.1c02728
• Publication Date: 2021-10-21T17:23:38Z
• Authors: Sourav Mishra; Jinseong Jeon; Jun-Kyu Kang; Sang-Hyun Song; Tae-You Kim; Changill Ban; Hayoung Choi; Yonggoo Kim; Myungshin Kim; Joon Won Park
• Keywords: 0301 basic medicine; 03 medical and health sciences; Neoplasms; Mutation; Biomarkers, Tumor; Humans; Point Mutation; Sensitivity and Specificity; Circulating Tumor DNA

Cell-free DNA (cfDNA) analysis, specifically circulating tumor DNA (ctDNA) analysis, provides enormous opportunities for noninvasive early assessment of cancers. To date, PCR-based methods have led this field. However, the limited sensitivity/specificity of PCR-based methods necessitates the search for new methods. Here, we describe a direct approach to detect KRAS G12D mutated genes in clinical ctDNA samples with the utmost LOD and sensitivity/specificity. In this study, MutS protein was immobilized on the tip of an atomic force microscope (AFM), and the protein sensed the mismatched sites of the duplex formed between the capture probe on the surface and mutated DNA. A noteworthy LOD (3 copies, 0.006% allele frequency) was achieved, along with superb sensitivity/specificity (100%/100%). These observations demonstrate that force-based AFM, in combination with the protein found in nature and properly designed capture probes/blockers, represents an exciting new avenue for ctDNA analysis.