Bacterial adhesion and subsequent biofilm formation on biomedical implants and devices are a major cause of their failure. As systemic antibiotic treatment is often ineffective, there is an urgent need for antimicrobial biomaterials and coatings. Biomaterials equipped with antimicrobial surface designs based on different mechanisms of action require different in vitro evaluation methods. Current industrial evaluation standard tests do not sufficiently account for different, advanced antimicrobial surface designs, yet are urgently needed to obtain convincing in vitro data for approval of animal experiments and clinical trials. The present study aims to provide innovative and clear guidance to choose appropriate evaluation methods for the three distinctly different mechanisms of antimicrobial design: (1) antimicrobial-releasing, (2) contact-killing and (3) non-adhesivity. Use of antimicrobial evaluation methods and definition of industrial standard tests, tailored toward the antimicrobial mechanism of the design, as identified here, fulfill a missing link in the translation of novel antimicrobial surface designs to clinical use. The aim of this study is to categorize presently available methods including industrial standard tests for the in vitro evaluation of antimicrobial surface designs according to their suitability with respect to their antimicrobial mechanism of action. Critical consideration of methods to evaluate antimicrobial efficacy clearly relates the different methods to a specific mechanism of antimicrobial action. The overview of methods presented will be illustrated with some recent coating developments, in particular the design of multi-functional coatings and contact killing designs. In addition to addressing differences and similarities between methods, attention will be given to microbial inoculum size, choice of species and the application of conditioning films. In an era in which animal experiments become increasingly difficult to obtain permission for, standardization is extremely important and may reduce the need for animal experiments.