Teeth offer nonshedding surfaces on which a wide range of bacterial species accumulate as thick, cohesive plaques. Intergeneric coaggregation mediated by specific recognition between surface "cohesins" is thought to contribute to both the cohesiveness of plaque and the sequence in which bacteria colonize the tooth surface. There is some evidence that Gram-positive species, like the efficient tooth colonizer Actinomyces viscosus, enhance subsequent tooth colonization by the more virulent periodontal pathogen Bacteroides gingivalis. To study their mechanism of cohesion, we have developed an in vitro assay that measures the sequential binding of tritium-labeled B. gingivalis to A. viscosus adsorbed to saliva-coated hydroxyapatite beads, mimicking teeth (actinobeads). The assay yields equilibrium and kinetics data amenable to statistical analysis. The presence of A. viscosus significantly increased the number of B. gingivalis cells bound. Inhibition studies were conducted to test the sensitivity of binding to heat; to various saccharides and sugar amines; to proteolytic treatment of Bacteroides; and to incorporation of various chaotropic agents, increased KCl, and saliva in the suspension buffer. Heating the Bacteroides cells but not the actinobeads diminished Bacteroides adherence. Proteolysis and various saccharides had little, if any, effect. Among chaotropic agents, NaSCN and LiCl reduced numbers of cells bound by 40%, but tetramethylurea had no effect. Increasing the ionic concentration of KCl reduced binding by 50 to 60%. Diluted saliva showed a concentration-dependent inhibition of B. gingivalis adherence to actinobeads. To begin examining B. gingivalis surface molecules significant to these reactions, lipopolysaccharide was extracted by the phenol–water method and analyzed by biochemical assays and polyacrylamide gel electrophoresis. Both homologous and heterologous lipopolysaccharides partially inhibited binding in a concentration-dependent manner, but actual binding of B. gingivalis lipopolysaccharide to A. viscosus cells could not be demonstrated. While heat, lipopolysaccharide, chaotropic agents, and concentrated KCl interfered to a limited extent with B. gingivalis – A. viscosus interactions on a surface mimicking teeth, the actual mechanism of cohesion is currently obscure and under investigation. The relevance of in vitro coaggregation experiments to colonization by periodontal and other pathogens was discussed.