Saliva is increasingly used for diagnostic purposes, but little is known about the potential effect of toothbrushing on the composition of saliva. Therefore, saliva collected before and 0, 15, 30 and 45 min after toothbrushing according to the Bass method was analysed for relevant protein composition. The flow rate initially increased (+15%) after toothbrushing, followed by a decrease after 15 min (-15%). The concentration of salivary immunoglobulin A (S-IgA) decreased (-40%) whereas the albumin concentration increased (+400%) within 20 min. Flow rate and S-IgA concentration returned to baseline values within 30 min. The albumin concentration, however, was still increased after 45 min. No significant increases were observed in the concentrations of total protein and amylase. Our results show that toothbrushing has a significant effect on the protein composition of saliva, by contamination with serum constituents. This should be taken into account when saliva is used for diagnostic purposes.