Abstract
A surface-enhanced Raman scattering (SERS)-based aptasensor was developed for the rapid and sensitive detection of Escherichia coli (E. coli), a major pathogen in urinary tract infections (UTIs). The sensor utilizes magnetic beads embedded with gold nanoparticles (MB-AuNPs) functionalized with capture DNA (cDNA) as both the SERS-active substrate and magnetic separation tool. The detection mechanism relies on an aptamer DNA-probe DNA complex: when the aptamer binds specifically to E. coli, the probe DNA is released and subsequently hybridizes with cDNA on the MB-AuNPs. This brings a Cy5 Raman label close to the gold surface, generating a strong SERS signal. The assay offers a one-step process, eliminating the need for bacterial culture or nucleic acid amplification, and completes within approximately 6 h. Quantitative analysis demonstrated a detection limit of 5.9 × 10(3) CFU/mL, well below the clinical threshold for UTIs, with a reliable calibration curve (R(2) = 0.990). Selectivity tests confirmed high specificity for E. coli without cross-reactivity to other bacteria. Clinical evaluation using 21 urine samples showed high diagnostic performance: 100% sensitivity, 91% specificity, 95% accuracy, and 100% precision compared to standard urine culture. These results highlight the aptasensor's potential as a rapid, sensitive, and specific alternative for UTI diagnosis in clinical settings.