A novel DNA biosensor for the ultrasensitive detection of DNA methyltransferase activity based on a high-density "hot spot" SERS substrate and rolling circle amplification strategy

• Published in: Analyst (London) Vol. 146; no. 17; pp. 5326 - 5336
• Main Authors: Ge, Shengjie, Ran, Menglin, Mao, Yu, Sun, Yue, Zhou, Xinyu, Li, Li, Cao, Xiaowei
• Format: Journal Article
• Language: English
• Published: London Royal Society of Chemistry 07.09.2021
• Subjects: Amplification; Arrays; Biosensors; Cancer; Density; Gold; Nanoparticles; Reliability analysis; Silicon dioxide; Substrates
• DOI: 10.1039/d1an01034d
• ISSN: 0003-2654; 1364-5528

Herein, we proposed a novel biosensor based on a high-density "hot spot" Au@SiO 2 array substrate and rolling circle amplification (RCA) strategy for the ultrasensitive detection of CpG methyltransferase (M.SssI) activity. In the presence of M.SssI, the RCA process can be triggered, causing the augmentation of the single-stranded DNA (ssDNA) at the tail of the double-stranded DNA (dsDNA), and the ssDNA can be hybridized with numerous DNA probes labeled with Raman reporters in the next steps. Afterwards, the resultant ssDNA can be modified to the Au@SiO 2 array substrate with the SERS enhancement factor of 7.49 × 10 6 . The substrate was synthesized by using a monolayer SiO 2 array to pick up the Au nanoparticle (AuNP) array and finite-difference time-domain (FDTD) simulation showed its excellent SERS effect. Particularly, the developed biosensor displayed a significant sensitivity with a broad detection range covering from 0.005 to 50 U mL −1 , and the limits of detection (LODs) in PBS buffer and human serum were 2.37 × 10 −4 U mL −1 and 2.51 × 10 −4 U mL −1 , respectively. Finally, in order to verify the feasibility of its clinical application, the serum samples of healthy subjects and breast cancer, prostate cancer, gastric cancer and cervical cancer patients were analyzed, and the reliability of the results was also confirmed by western blot (WB) experiments. Taking advantage of these merits, the proposed biosensor can be a very promising alternative tool for the detection of M.SssI activity, which is of vital importance in the early detection and prevention of tumors. Herein, a novel biosensor based on a high-density "hot spot" Au@SiO 2 array SERS substrate and rolling circle amplification (RCA) strategy for the ultrasensitive detection of CpG methyltransferase (M.SssI) activity was devoloped.