Sensitive detection of miR-21 and miR-25 in gastric adenocarcinoma patient serum using a SERS sensor based on AuNT and enzyme cleavage strategy

Abstract

MicroRNA (miRNA) detection has significant application value for early cancer diagnosis. In this study, a surface-enhanced Raman scattering (SERS) sensor was developed for detecting miR-21 and miR-25 in the serum of Gastric adenocarcinoma (GAC) patients. The sensor was constructed using arrays of Au trioctahedral nanoparticles (AuNT) and enzyme cleavage techniques. The AuNT was obtained by self-assembly at the oil–water interface, and the Cy5-labeled miR-21 and 5-FAM-labeled miR-25 complementary single-stranded ssDNA-21 and ssDNA-25 were connected with the AuNT to form the SERS sensor. When miR-21 and miR-25 were present, ssDNA-21 and ssDNA-25 were paired and hybridized to form miR-21-ssDNA-21 and miR-25-ssDNA-25 double strands. Duplex-specific nuclease (DSN) could act on the DNA phosphodiester bond in the double strand, causing Cy5 and 5-FAM to be far away from the AuNT, which resulted in a reduction of the SERS signal. In the range of 10 aM to 1 pM, the logarithm of miR-25 concentration was linearly related to the intensity of the characteristic peak of 5-FAM at 1178 cm⁻¹, and the limit of detection (LOD) was determined to be 8.12 aM. The logarithm of miR-21 concentration was linearly related to the characteristic peak intensity of Cy5 at 1367 cm⁻¹, and the LOD was determined to be 4.29 aM. Furthermore, the accuracy of the SERS sensor for the detection of miR-21 and miR-25 in clinical serum samples was evaluated using real-time quantitative polynucleotide chain reaction (qRT-PCR) technology as the gold standard. The relative errors of the two methods miR-21 in healthy people and gastric adenocarcinoma patients were 1.71% and −2.40%. The relative errors of miR-25 were 2.74% and −2.67%. There was no significant difference between the two methods, and the expression levels of miR-21 and miR-25 in the serum of GAC patients were found to be higher than those in healthy individuals. Consequently, this method offers a reliable solution for the early diagnosis of gastric cancer.