A Monolithic, Thiol-Functionalized Au-Based Bio-CMOS Aptasensor for Rapid, Label-Free Detection of Escherichia coli...

Rapid, point-of-care detection of Escherichia coli O157:H7 remains an unmet clinical need, as culture and molecular methods are slow and poorly suited to decentralized or emergency settings. A label-free, monolithic aptasensor biochip was fabricated in a standard 65 nm CMOS process, featuring three aptamer-functionalized gold sensing pads with matched reference pads for differential readout. A 37-mer DNA aptamer targeting the E. coli O157:H7 lipopolysaccharide was immobilized via thiol-gold self-assembled monolayer chemistry. Binding events were transduced into surface-potential shifts, amplified by an on-chip analog front-end (~100 V/V gain, 101.5 µW), and evaluated using calibration standards, patient specimens, and hospital environmental samples, with fluorescence microscopy for validation. The sensor achieved 47.42 mV/decade sensitivity across 1-10,000 CFU/mL, an IUPAC detection limit near 3.74 CFU/mL, and an empirical LOD of about 11 CFU/mL, with outputs tracking bacterial load and ~5.7% matrix-related deviation. Hospital samples were detectable to 28 CFU/mL. Because the patient-derived and hospital-acquired cohorts (n = 10 and n = 6, respectively) were assembled for pilot analytical and matrix-tolerance characterization rather than for diagnostic-accuracy determination, these results establish detectability and matrix robustness in real clinical and environmental specimens rather than clinical diagnostic sensitivity or specificity, which will require a larger, prospectively enrolled cohort in future work. Sensor kinetics followed Langmuir-type adsorption, saturating within 16-25 min for target pathogens versus slower responses for non-target strains. Selectivity tests against six bacterial species showed discrimination, with cross-reactivity decreasing from related E. coli pathotypes to Enterobacteriaceae to Gram-positive species. Inter-pad variability stayed below 1.5 mV, supporting this compact, low-power platform for scalable, enrichment-free point-of-care pathogen detection.