Tooth stainingwith the use of chlorhexidine preparations is the major problem of long term application. Evidence suggests that the staining arises from a cationic/anionic interaction of chlorhexidine with components of certain dietary materials. The purpose of thisin vitrostudy was to compare visually the development of tea and coffee staining on acrylic and tooth specimens treated with chlorhexidine and to follow the development of tea staining on perspex by light microscopy and color television image analysis. All specimens were maintained in their respective beverage for 5 days with test specimens removed three times a day and placed for 2 minutes in an 0.2% chlorhexidine solution. Both test tooth and acrylic specimens showed comparable and markedly increased staining by the beverages compared with control specimens. Color television image analysis of test specimens demonstrated more marked and rapid development of tea staining when studied on a daily basis. Microscopic examination revealed the staining to be made up of small particles of material which increased in size and coalesced with time. Again, marked differences were apparent in the stain on test and control specimens. The results of thisin vitromethod provided further evidence for a dietary aetiology to chlorhexidine staining and were consistent with clinical findings. Such a method may be useful to assess staining arising from the use of other anti‐plaque agents.