This review gives a survey on the state of the art of pesticide detection usingflow-based biosensing systems for sample screening. Although immunosensor systems havebeen proposed as powerful pesticide monitoring tools, this review is mainly focused onenzyme-based biosensors, as they are the most commonly employed when using a flowsystem. Among the different detection methods able to be integrated into flow-injectionanalysis (FIA) systems, the electrochemical ones will be treated in more detail, due to theirhigh sensitivity, simple sample pretreatment, easy operational procedures and real-timedetection. During the last decade, new trends have been emerging in order to increase theenzyme stability, the sensitivity and selectivity of the measurements, and to lower thedetection limits. These approaches are based on (i) the design of novel matrices for enzymeimmobilisation, (ii) new manifold configurations of the FIA system, sometimes includingminiaturisation or lab-on-chip protocols thanks to micromachining technology, (iii) the useof cholinesterase enzymes either from various commercial sources or genetically modifiedwith the aim of being more sensitive, (iv) the incorporation of other highly specificenzymes, such as organophosphate hydrolase (OPH) or parathion hydrolase (PH) and (v) thecombination of different electrochemical methods of detection. This article discusses thesenovel strategies and their advantages and limitations.