If a fermentation process is followed by a change in the concentration of maltose, information on the level of the maltose is important. A new type of sensor was developed for measuring the content of maltose in fermentation broth. The base of the sensor is a thin-layer reactor, in which a protein membrane was chosen for the immobilization of enzymes. To measure maltose, we investigated the influence of enzymes such as α-glucosidase (EC 3.2.1.20) any amyloglucosidase (EC 3.2.1.3) on the effectiveness of conversion to glucose. Because amyloglucosidase proved to be more effective, a mixture of amyloglucosidase and glucose oxidase was applied for the determination of maltose. To develop the best measuring technique, the consequences of changes in different parameters, such as the optimal ratio of enzymes, role of pH value and that of the flow rate, were studied. An amperometric measuring cell with PtAg/AgClPt electrodes was used at +600 mV operating potential. The results indicate that the maltose content in different types of fermentation broth can be determined by the new measuring cell in the range of 0·2–4 m m maltose. The cell was successfully tested in the fermentation of brewer's yeast.