A label-free immunosensor for the direct detection of cholera toxin (CT) at sub-attomolar level has been developed based on potential-step capacitance measurements. Anti-CT antibody was adsorbed on gold nanoparticles (AuNPs) incorporated on a polytyramine-modified gold electrode. The concentration of CT was determined by detecting the change of capacitance caused by the formation of antibody-antigen complexes. By using AuNPs adsorbed to the sensing surface, the signal was dramatically increased leading to a significantly more sensitive assay. In fact, under optimum conditions the immunosensor could detect CT concentration with a limit of detection of 9 x 10(-20)M or 0.09 aM, with a dynamic range between 0.1 aM and 10 pM. Good analytical reproducibility could be obtained by injecting CT up to 36 times with an RSD of 2.5%. In addition, good performance of the developed immunosensor was achieved when applied to turbid water samples collected from a local stream that were spiked with CT.