Background: Evidence to date shows that mast cells play a critical role in immune defenses against infectious agents, but there have no reports about involvement of these cells in eliminating periodontopathogens. In this study, the phagocytic ability of mast cells against A. actinomycetemcomitans in comparison with macrophages was evaluated. Methods: In vitro phagocytic assays were conducted using murine mast cells and macrophages, incubated with A. actinomycetemcomitans, either opsonized or not, with different bacterial load ratios. After one hour, cells were stained with acridine orange and assessed by confocal laser scanning electron microscopy. Results: Phagocytic ability of murine mast cells against A. actinomycetemcomitans was confirmed. In addition, the percentage of mast cells with internalized bacteria in absence of opsonization was higher than those in presence of opsonization. Both cell types showed significant phagocytic activity against A. actinomycetemcomitans. However, the percentage of mast cells with non-opsonized bacteria was higher than those of macrophages with opsonized bacteria, in one of ratios (1:10). Conclusion: This is the first report about the participation of murine mast cells as phagocytes against A. actinomycetemcomitans, mainly in the absence of opsonization with human serum. Our results may indicate that mast cells act as professional phagocytes in the pathogenesis of biofilm-associated periodontal disease.