A sensitive method for the detection of staphylococcal enterotoxin B (SEB) using a flow-injection capacitive biosensor is presented. SEB was purified from a crude culture filtrate of Staphylococcus aureus through three chromatographic steps. The first two steps were based on ion-exchange chromatography, and the last step was carried out on a gel filtration column. The SEB recovery values after the purification stages were 88%, 74%, and 12%, respectively. A horseradish peroxidase labeled antistaphylococcal enterotoxin B was prepared by the periodate method and was further employed in a sandwich-enzyme-linked immunosorbent assay (ELISA) for the determination of SEB concentrations in different samples obtained during the processing of the crude filtrate. The capacitive biosensor could detect SEB concentrations as low as 0.3 pg ml(-1) with a linearity ranging from 2.8 pg ml(-1) to 2.8 ng ml(-1) under optimized conditions. The response time was about 10 min. A good agreement was achieved between the developed capacitive biosensor system and ELISA as a reference method for detection of SEB levels in different purification samples. The newly developed sensor has the benefits of simplicity, high sensitivity, and multiple use capability.