A major concern related to the adoption of genetically modified (GM) crops in agricultural systems is the possibility of unwanted GM inputs into non-GM crop production systems. Given the increasing commercial cultivation of GM crops in the European Union (EU), there is an urgent need to define measures to prevent mixing of GM with non-GM products during crop production. Cross-fertilization is one of the various mechanisms that could lead to GM-inputs into non-GM crop systems. Isolation distances between GM and non-GM fields are widely accepted to be an effective measure to reduce these inputs. However, the question of adequate isolation distances between GM and non-GM maize is still subject of controversy both amongst scientists and regulators. As several European countries have proposed largely differing isolation distances for maize ranging from 25 to 800 m, there is a need for scientific criteria when using cross-fertilization data of maize to define isolation distances between GM and non-GM maize. We have reviewed existing cross-fertilization studies in maize, established relevant criteria for the evaluation of these studies and applied these criteria to define science-based isolation distances. To keep GM-inputs in the final product well below the 0.9% threshold defined by the EU, isolation distances of 20 m for silage and 50 m for grain maize, respectively, are proposed. An evaluation using statistical data on maize acreage and an aerial photographs assessment of a typical agricultural landscape by means of Geographic Information Systems (GIS) showed that spatial resources would allow applying the defined isolation distances for the cultivation of GM maize in the majority of the cases under actual Swiss agricultural conditions. The here developed approach, using defined criteria to consider the agricultural context of maize cultivation, may be of assistance for the analysis of cross-fertilization data in other countries.