Recombinant antibodies provide an emerging strategy in the development of new immunosensors. In particular, single chain antibodies (scFvs) can be isolated and expressed in bacterial systems that also allow their in vitro manipulation at the gene level. In this work, we present for the first time results of single-chain phage displayed antibodies combined with amperometric detection and its application as an immunosensor. The scFv is immobilized on a carbon electrode and used to capture and quantify its specific target antigen. We describe the detection of the sugar milk lactose, the bacteria Listeria monocytogenes, and the enzyme MtKatG, which is expressed by Mycobacteriumtuberculosis.