A new lactate sensor makes it possible to measure the lactate content of whole blood directly in less than 1 min, using only a 10-microL blood sample. The procedure works equally well with plasma, serum, spinal fluid, other body fluids, or tissue homogenates. The instrument is calibrated with lactate standards between 0 and 15 mMol/L. The sensor, a polarographic enzyme electrode, gives a current which is a linear function of the lactate concentration. There is no interference from glucose, pyruvate, alcohol, ascorbate, anticoagulants, lidocaine, acetaminophen, or other drugs and metabolites commonly encountered in critically ill patients. The lactate sensor is composed of a peroxide sensor and an enzyme transducer membrane. The lactate is stoichiometrically converted to pyruvate and hydrogen peroxide by lactate oxygen oxidoreductase derived from Pediococcus species. The oxygen required for the enzymatic oxidation is supplied via an air-permeable silicone elastomeric membrane used for stirring. Comparison of our new electroenzymatic method with the Boehringer-Mannheim photoenzymatic method gives correlations of 0.997 for both whole blood and plasma.