Bound sialic acid (b-SIA) occurs linked to macromolecules in organisms. An electrochemical method for determining b-SIA, using a potentiometric four-channel thick-film sensor, has been developed. The sialidase sensor consists of a bilayer of a membrane containing sialidase isolated from Clostridium perfringens immobilized in a poly(vinyl acetate)–polyethylene copolymer, which is then placed on top of an H⁺-selective poly(vinyl chloride)–poly(vinyl acetate) indicator membrane. The enzymic cleavage of the linkage to the macromolecular substrate leads to the release of sialic acid, which is an α-ketocarboxylic acid with a pK_a of 2.6. The decrease in the pK_a value of the carboxyl group results in a local pH change inside the sialidase containing sensor membrane, which is monitored by the H⁺-selective indicator membrane. The determination of non-pathological (0.5–0.8 mg cm^–3) and pathological levels (1.3–3.0 mg cm^–3) of b-SIA in serum samples prepared from human test sera is shown. The detection limit was found to be 0.3 mg cm^–3 of b-SIA in these sera, and the linear range was from 1.0 mg cm^–3 to more than 10.0 mg cm^–3 of b-SIA. The slope of the sensor response was found to be dependent on the relative molecular mass of the substrates, which is crucial for quantitative measurements. The pH optimum of the sialidase sensor was found to be pH 4.0 for the substrates sialyl-lactose, mucine and colominic acid. The serum samples were diluted with an equal volume of 100 mmol dm^–3 phosphate buffer in order to bring the final pH to 4 before analysis. The rise time to reach 90% of the final reading ranges from 2 to 5 min for the different substrates. The slope of the sensor response decreases to 55% of the initial value during 100 h of continuous operation at room temperature at pH 4.0. By averaging the readings from the four channels of the sensor the accuracy for the determination of b-SIA in human sera can be improved and a standard deviation of ±2.5% is achieved.