Chemometric theory was applied to a microbial sensor for determinations of ethanol in the presence of glucose. Microbial sensors, consisting of Gluconobacter oxydans cells immobilized on Clark-type amperometric oxygen electrodes, exhibited good sensitivity but low selectivity toward ethanol and glucose. An Eksan-G commercial glucose analyzer was used as a second sensor for multivariate calibration and analyses. Microbial sensors exhibited nearly complete additivity for total glucose plus ethanol concentrations from 0.0 to 0.6 mM. Within this linear range, chemometric analyses provided estimates of ethanol concentration with measurement errors of less than 8%. Multivariate calibration thus is a promising approach to enhance the usefulness of microbial sensors.