Plaque formation on buccal tooth surfaces was studied by replica technique, consisting of impressions using low viscosity silicone impression materials and positive models produced in epoxy resins. Bacterial accumulation occurred near the cemento-enamel junction in 6-hr specimens, and subsequently expanded in a coronal direction. This development took place partly by extensions of single layers of bacteria, and partly by a pattern where the colonization was mostly restricted to vertical enamel cracks. Plaque accumulations were also frequently located in abrasion grooves and surface pits in the enamel, and prolific plaque areas were consistently surrounded by a monolayer of bacterial cells. Globular and hemispheric structures which occurred, especially on root surfaces immediately after cleaning, were probably artefacts caused by air bubbles or remaining moisture. In separate series of experiments it was demonstrated that improved reproduction of details from the plaque could be achieved by repeating the replicating procedure. The findings indicate that plaque formation starts by adsorption and proliferation of individual bacteria on tooth surfaces, and not by adsorption of aggregates of cells. Special attention should be directed against the problem of artefacts and moisture in replica studies of dental plaque.