An amperometric phosphate biosensor, based on a cobalt phthalocyanine screen-printed carbon electrode (CoPC-SPCE) is described. The immobilisation of the enzyme pyruvate oxidase (PyOd) was investigated using pre-formed cellulose acetate/cellulose nitrate membranes, of different pore sizes, and the cross-linking agent, glutaraldehyde (GLA). The latter method was found to be superior in terms of performance characteristics and also ease of fabrication. A linear range of 2.5-130 M and limit of detection of 2 M was obtained under optimal conditions. The biosensor also exhibited an excellent response time of just 13 s 1, n = 3. The biosensor was successfully applied to the measurement of phosphate in pond water samples; the mean recovery of spiked water samples was 103.2%, n = 3. The original concentration of phosphate calculated in the water sample (48 M) was found to be in good agreement with that found using a standard colourimetric method. In addition to the analysis of water samples the biosensor was also applied to the analysis of human urine, with only a simple dilution of the sample, directly into the electrochemical cell, required for analysis. The precision of the biosensors, obtained during the urine analysis was 6.4%, n = 6.