The purpose of the present study was to investigate whether functionally different macrophages are present in clinically healthy gingiva and during human experimental gingivitis. Eight male probands were introduced to an oral hygiene program until all reached mean Plaque and Gingival Index scores approaching zero. During the following 19 days all oral hygiene was abandoned. At d -14, 0, 2, 4, 7, 11 and 19 clinical indices and gingival biopsies were taken. Cryostat sections were incubated with monoclonal antibodies against mature macrophages (25F9), inflammatory macrophages (27E10) and anti-inflammatory macrophages (RM 3/1). Positive cells were counted in the inflammatory infiltrate (IF) and the connective tissue (CT). At d -14 elevated numbers of 27E10-positive cells were observed which decreased significantly at d 0 (p less than 0.018) and increased again at d 19 (p less than 0.026). Significant differences in the number of RM 3/1-positive cells were found between d 0 and d -14, 2, 4 and 7 (p less than 0.05) while no differences in the number of 25F9-positive cells were observed throughout this study. It was concluded that experimental gingival inflammation is characterized by the appearance and disappearance of functionally different macrophage subpopulations.