An amperometric procedure is described for the determination of glycerol and triglycerides in aqueous samples and in serum using glycerol dehydrogenase immobilized on a collagen membrane. Glycerol is determined by measurement of the steady-state oxidation currents generated at a platinum electrode by NADH produced in the enzyme-catalyzed reaction. The triglycerides were first hydrolyzed by the enzyme lipase in solution and the resulting glycerol determined similarly. Olive oil, determined to contain 78 % triolein, was used as the source of triglycerides in this study. For both glycerol and triglycerides the calibration plots are linear in the range from 0 to 12 μM, with detection limits of 0.2 and 0.7 μM, respectively. The immobilized glycerol dehydrogenase retained high operational activity for a period longer than 30 days.