An in vitro system for the analysis of changes in depth distribution of diffusates in bacterial films

No suitable technique exists which allows simultaneous analysis of time-and depth-dependent concentrations of all components of interest in single samples of intercellular fluid from dental plaque biofilms. We have developed an in vitro model which allows detailed study of these interactions by analysis of intercellular fluid and pH measurement at the film base. Compact, defined-depth films of Streptococcus mutans were formed in a micrometer-controlled, variable-depth well and bathed in synthetic saliva. Films exposed to synthetic saliva containing glucose for 2 min followed by clearance with glucose-free synthetic saliva gave typical 'Stephan-type' pH profiles. Intercellular fluid isolated from successive 200-mu m-thick sections of 600-mu m-deep films was analysed by ion chromatography. A concentration gradient of lactate, falling with depth, was measured. The experimental system described here, the first of its type to be described, can easily be used to analyse pH changes and the depth-dependent distribution of diffusates in a model bacterial film. Although this bacterial film is far removed from a natural biofilm, the apparatus has potential for the study of grown biofilms and is an important advance towards position-dependence analysis of diffusates in biofilms.