One of the most powerful techniques in molecular biology is the controlled expression of specific proteins by transfection of eukaryotic cells. This method has become feasible and highly sensitive and, thus, suitable for high-throughput reporter gene assays in basic and applied research. Moreover, the limiting factors are neither the transfection efficiency nor the functional analysis, but rather the ability to manage complex experimental protocols when multiple genes are co-transfected and/or when the effects of several chemical compounds are investigated within the same experiment. Here, we describe an easy-to-use and highly flexible spreadsheet template intended to rationalize and expedite the organization and data management of multi-step reporter gene assays. The objectives of this spreadsheet template are the design of the transfection protocol, the coordination of the administration of test compounds, and the graphical presentation and statistical analysis of the results.