AIM: To detect the presence of milk fat globule-epidermal growth factor-factor 8 (MFG-E8) in human gingival crevicular fluid (GCF) and to determine its potential role as a biomarker for periodontal disease activity. MATERIALS & METHODS: GCF was collected from a total of 230 sites from with seven subjects with gingivitis, twelve subjects with chronic moderate periodontitis, fourteen subjects with chronic severe periodontitis, six subjects with localized severe periodontitis as well as seven clinically healthy subjects. Subjects from the severe periodontitis group received nonsurgical therapy and were re-evaluated after 4 weeks. GCF was re-collected from thirty sites in five subjects at that point. Pocket reduction surgery was performed on the same subgroup and GCF was collected again at the 4-month postoperative appointment. GCF collection was performed using paper strips and analyzed for the presence of MFG-E8 and cytokines using multiplexing magnetic bead immunoassays (Luminex xMAP with MagPlex beads). Each sample was tested using an MFG- E8 kit, a human cytokine/chemokine kit and a human bone panel kit. RESULTS: MFG-E8 was detected at higher levels in sites with gingivitis and gingival health as compared to all periodontitis groups, suggesting that MFG-E8 production is down regulated in periodontitis. Consistent with this notion, MFG-E8 was found to significantly increase following non-surgical therapy of subjects with severe periodontitis. Furthermore, the levels of MFG-E8 significantly increased after surgical treatment correlating with decreased probing pocket depths. IL-1α, IL-1β, RANKL, OPG, IL-6 and IL-17A were detected at levels consistent with those found in earlier studies. In the periodontitis treatment subgroup, the levels of RANKL, IL-6 and IL-17A decreased with decreasing probing pocket depths. CONCLUSIONS: MFG-E8 was detected in human GCF collected from healthy, gingivitis and periodontitis subjects using a magnetic bead-based immunoassay. The levels of MFG-E8 were negatively related to the level of gingival inflammation and increased after both non-surgical and surgical treatment of periodontal disease. These data suggest the potential of MFG-E8 as a novel biomarker of periodontal disease.