Apart from the decisive sensitivity and specificity of immunosensors, the employed antibodies essentially contribute to additional key factors like fabrication costs for sensor chips and sensor stability. A production scheme for recombinant antibody fragments has been optimised with respect to these particular issues of biosensor development. The phagemid vector pCANTAB 5 E is widely used for the selection of antibody fragments from corresponding libraries. However, large-scale production of the selected single-chain F(v) (scFv) fragments is substantially restricted by the high cost for the inducer IPTG and the anti-E-tag antibody. The latter is needed in significant amounts for the purification of the recombinant protein. A generic strategy was established for subcloning scFv variable regions from pCANTAB 5 E into the plasmid pASK85 for the expression of F(ab) fragments. pASK85 bears coding sequences for murine constant domains including a His(6) tag at the carboxyl-terminal end of the constant heavy chain domain. The anti-s-triazine antibody K47H served as a model system in this study. Biosynthesis of the F(ab) fragment in a high cell density fermenter was induced by addition of anhydrotetracycline. The F(ab) fragment was subsequently purified from the periplasmic extract in a single step by immobilized metal affinity chromatography (IMAC). A yield of 100 microg/lxOD(550) purified F(ab) fragment was obtained employing a standard fermentation scheme. The sensitivity and cross-reactivity of the F(ab) was comparable to the parent scFv when assayed by enzyme immunoassay. However, the F(ab) fragment exhibited significantly improved long-term stability.