MXene Boosted Ultrasensitive Electrochemical Detection of 5‐Hydroxymethylcytosine in Genomic DNA from Complex Backgrounds

Abstract 5‐Hydroxymethylcytosine in genomic DNA (5hmC‐DNA), a predominant epigenetic mark, plays key roles in a broad range of bioprocesses such as cancer progression. However, efficient detection of 5hmC‐DNA, especially from highly complex biological specimens, remains a harsh challenge due to its low abundance as well as the serious disturbance of non‐targets. Leveraging MXene as the signal booster, an electrochemical biosensing technique is presented here for ultrasensitive detection of 5hmC‐DNA from complex biological samples. The developed technique integrates enzyme‐catalyzed transform of the hydroxyl group of 5hmC, sequence‐specific recognition of the 5hmC‐DNA targets, and facile electrochemical reaction. Remarkably, nanosized Ti₃C₂T_x MXene is introduced as a highly conductive agent, managing to greatly boost the signals. Exhibiting outstanding consistency and robustness, the technique not only achieves excellent linear responses in a broad range of 5hmC‐DNA (1.0 × 10⁻¹³–1.0 × 10⁻⁹ m) with the lowest detection limit of 59 fM but also possesses excellent selectivity against 5hmC analogs of including cytosine (C)‐ and 5‐methylcytosine (5mC). More importantly, a proof‐of‐concept validation demonstrates the extraordinary ability of this technique for directly detecting 5hmC‐DNA from complex backgrounds including cellular genomic extracts and serum of irradiated or non‐irradiated mice, suggesting its highly promising potential for diverse applications such as early cancer screening.