The purpose of this study is to evaluate biofilm formation by the bacteria involved in chronic rhinosinusitis. Mixed cultures of the pathogens in chronic rhinosinusitis including coagulase negative Staphylococcus epidermidis, Hemophilus influenzae, Pseudomonas aeruginosa, Moraxella catarrhalis and Streptococcus pneumoniae were obtained. The microbial attachment and biofilm formation was measured by a crystal violet based microtitre assay. In order to grow biofilms under flowing conditions, a CDC laboratory biofilm reactor system was used. Pure, then mixed species biofilms were examined as a preliminary study for a larger project in which innovative treatments will be tested in vitro. P. aeruginosa has the maximum biofilm forming capacity. This pathogen showed a steady growth rate in pure species as well as in mixed species biofilm formation. The pure culture of S. epidermidis was able to grow biofilms, but it decreased in numbers from the initial level when it was combined with P. aeruginosa. M. catarrhalis, also has the capacity to form biofilms, but showed an increased biofilm cell density when it was grown in mixed culture. S. pneumoniae and H. influenzae were not good biofilm formers in pure culture, but these pathogens showed some attachment when they were combined in mixed culture. The crystal violet based microtitre assay and the CDC laboratory biofilm reactor system, are suitable for in vitro study, but represents a very artificial condition. The next stage is to study mixed films in a flowing system and ultimately on sinus tissues.