Ag + /Hg 2+ -Controlled Programmable DNA Nanostructures Enable Tumor Liquid Biopsy by Mass Spectrometry Simultaneous Sensing of Multiple Circulating Tumor DNAs

Circulating tumor DNA (ctDNA) has emerged as a critical noninvasive biomarker for cancer diagnosis and monitoring. To effectively capture tumor heterogeneity, we developed a rapid, homogeneous, and label-free method for the simultaneous detection of EGFR L858R and 19 del mutations in lung cancer, employing a filter-membrane-assisted separation strategy coupled with inductively coupled plasma mass spectrometry (ICP-MS). The assay utilized functional Y-shaped DNA nanospheres, self-assembled through T-Hg 2+ -T and C–Ag + -C coordination, enabling highly specific target recognition and efficient release of metal reporters. Following filtration, free Hg 2+ and Ag + ions were quantitatively measured by ICP-MS, facilitating multiplexed ctDNA detection with high sensitivity. The method demonstrated a wide linear range from 10 aM to 10 pM, with detection limits as low as 4.3 aM for L858R and 3.7 aM for 19 del. Clinical validation using 42 plasma samples revealed 100% specificity and 96.9% sensitivity, closely aligning with the pathological findings. Receiver operating characteristic (ROC) analysis achieved an area under the curve (AUC) of 0.94, highlighting the excellent diagnostic accuracy of this approach. This strategy provides a robust, noninvasive platform for precise lung cancer diagnosis and treatment monitoring.