Moraxella catarrhalis is a major cause of infectious exacerbations of chronic obstructive lung disease. Adhesion of this pathogen to epithelial cells is critical for its pathogenicity. Although much work has been done on identifying surface molecules of M. catarrhalis as adhesins, several adhesion assays were used in these studies which has never been validated or compared to each other. In the present study, we have examined the capacity of M. catarrhalis to adhere to different human epithelial cells. By using the two most commonly used adhesion assays based on the enumeration of colony-forming units or on the counting of adherent bacteria per epithelial cell by light microscopy, we identified significant limitations of both methods. These arose either from differences in strain-specific adhesion pattern on the epithelial cell surface or the dependence on the state of confluence of the epithelial cell layer. We developed a new fluorescence-based adhesion assay and compared our results to the two conventional methods. We demonstrated that the fluorescence-based adhesion assay offers a reliable and convenient method for the quantification of M. catarrhalis adhesion to confluent epithelial cell monolayers.