B-cells extracted from periodontal disease tissue were analyzed for the presence of activation markers using a range of monoclonal antibodies. In adult periodontitis (AP), 6% of B-cells expressed the IL-2 receptor (CD25) compared with 1-2% in peripheral blood and healthy or marginal gingivitis (H/MG) gingival B-cells. There was also an increase in the mean percentage of IgD-positive B-cells and a decrease in CD21 and CD22 expression. In both AP and H/MG lesions, 20-22% of the B-cells expressed CD23 compared with less than 5% in peripheral blood. As B-cells are activated by day 3 in culture and start differentiating into immunoglobulin-secreting cells by day 6, B-cell phenotypes were assayed at these times in this study. Following stimulation with the periodontopathic bacterium Porphyromonas gingivalis, the expression of CD23, CD21 and CD22 on B-cells extracted from AP lesions remained relatively constant over the 6-d culture period. However, with Fusobacterium nucleatum stimulation, there was a significant decrease in CD23, CD21 and CD22 expression after 3 d in culture, which corresponds to the activation time for B-cells. These results show that B-cells extracted from periodontal disease tissue display a range of activation markers and on stimulation, demonstrate differing responses to individual periodontopathic bacteria.