Afatoxins are toxic natural substances and one of the most prevalent types of mycotoxin contamination. They represent a high risk to human health as they contaminate a variety of agricultural raw materials. National and international institutions and organisations such as the European Commission have set regulatory limits for major mycotoxin classes. Several diferent methods for the detection and quantifcation of afatoxins have already been developed. On the one hand, highly sophisticated analytical reference methods like liquid chromatography coupled with mass spectrometry are applied, on the other hand, rapid screening methods based on immunochemical techniques have also been developed. Immunoassays are based on antigen-antibody interactions but the use of antibodies has many restrictions such as a low antibody stability and high costs of production. In recent years, an innovative method based on a new class of molecules, named aptamers, has been developed for analytical applications. Aptamers are specifc oligonucleotides which are able to bind to a target substance with high affnity and specifcity due to their three-dimensional, specifc structures. Among the methodologies used for transforming aptamer-target interactions into a visible signal, the horseradish peroxidase-like DNAzyme has been used frequently to catalyse chemical reactions to amplify biosensing events. DNAzymes are catalytic nucleic acids that act like enzymes and therefore represent good alternatives to enzymes. Like the horseradish peroxidase, the HRP-mimicking DNAzyme can also catalyse the oxidation of 3,3',5,5'-Thetramethylbenzidine (TMB) by the use of H2O2 as oxidising agent to produce a visible signal. In the overall project, research is done on an aptamer-based assay for the detection of a-atoxin B1 which should allow the rapid detection of this mycotoxin in maize. In this work, several factors of the DNAzyme-catalysed H2O2-mediated oxidation of TMB including the DNA sequence of the DNAzyme, substrate buffer, concentrations of TMB, H2O2 and hemin as well as the concentrations ratios and reaction time have been investigated and gradually optimised for this sensor developing purposes. The use of higher concentrated substrate and stopping solution and simultaneously less reaction volume was found to lead to higher absorbance values. Furthermore, the best results for sensor-developing purposes could be achieved when having a concentration ratio of TMB and H2O2 of 1:3 and DNAzyme and hemin of 1:8. Among the different substrate buffers tested, the most consistent results in the absorbance measurements with the lowest error rates could be achieved by using a substrate buffer containing 300 mM citric acid and 1 mM potassium sorbate. Moreover, acetonitrile was found to have the ability to enhance the catalytic reaction leading to higher absorbance values. Due to the optimisation of these factors, we were able to achieve a 50-times increase in sensitivity of the DNAzyme-catalysed H2O2-mediated oxidation of TMB compared to the beginning of the optimisation of this assay.