A novel application of a hybrid biosensor is here employed as an analytical method for the detection and presumptive identification of beta-lactam residues in milk. The method is based on measurements of carbon dioxide (CO2), the production of which is related to the microbial growth of the test microorganism Bacillus stearothermophilus var. calidolactis. The presence of beta-lactams in milk inhibits microbial growth and, consequently, the CO2 production rate. The analysis is based on the variation of CO2 between a milk sample spiked with beta-lactams and a twin milk sample containing beta-lactams plus a broad spectrum beta-lactamase, using an electrochemical device of biosensor. A blank milk sample is included as control. The result is obtained starting from the first 120 min. Moreover, the ability to recognize all of the beta-lactams speeds the total time of analysis when chemical identification and quantification are required. The analytical method appears to be adequate for milk control for qualitative screening purposes, complying with the requirements stated in Decision 2002/657/EC.