A Label-Free Cell-Based Biosensor Method for Ethanol Quantification Using Temperature-Induced Spontaneous Cell Detachment

Rapid, low-cost ethanol quantification is vital for beverage quality control, biofuel production, and pharmaceutical applications, yet current approaches are costly, reagent- or label-dependent, or rely on spectroscopy with substantial sample preparation. We introduce a purely cell-based, label-free biosensor that exploits temperature-gradient-induced spontaneous detachment of Saccharomyces cerevisiae from a chip surface. The readout is the detachment half-time, td50, derived from time-resolved changes in interfacial thermal resistance, Rth, at the solid–liquid interface. Cells were pre-exposed to ethanol (0–70% v/v) and the detachment kinetics monitored using the heat transfer method (HTM). Under these conditions, cells display a pronounced non-monotonic td50 response with a peak around 20% v/v ethanol. Overall, the td50 rises from ~45 min (0% ethanol) to ≳10 h (20%) and then decreases, with no detachment at 60% and beyond. Critically, cell quality gates the detachment window. Fresh yeast responds up to ~50%, whereas aged yeast ceases to detach above ~8%, demonstrating a dual-function assay. Complementary measurements show that ethanol decreases surface tension monotonically, as expected, while optical/SEM imaging reveals aggregation above the detachment window. Requiring only a heater and a temperature probe, this platform offers a compact and low-cost strategy for ethanol sensing. Its applicability in a complex matrix is further demonstrated using whiskey diluted to selected alcohol concentrations, which produced responses consistent with the ethanol calibration trend. Potentially, it also offers a thermal assay for real-time monitoring of microbial cell quality across biotechnology and bioengineering applications. Considering ethanol as a proxy for drugs, the strategy may also support label-free drug screening on cells. At a fundamental level, the non-monotonic effect of ethanol, and especially the sharp maximum at 20%, remains unresolved and invites further studies.