Background Ki-67, a nuclear protein is found in cells undergoing proliferation during the cell cycle. It has been established as an important tumor proliferation and prognostic marker. An increased expression of Ki-67 is observed in inflammation. Periodontitis tissue biopsy specimens have shown Ki-67 expression. Saliva has become popular as a non-invasive source of biomarkers that may have a clinical utility. Ki-67 has not been examined in the saliva of patients having periodontitis. This cross-sectional study aimed to detect and make a comparative estimation of salivary Ki-67 in health and periodontitis. Methods Fifty-two participants were divided equally into two groups: Health [systemically and periodontally healthy (n = 26)] and Periodontitis [systemically healthy periodontitis patients (n = 26)]. Study volunteers were recruited based on the selection criteria. Plaque index, modified gingival index, bleeding on probing, probing pocket depth and clinical attachment loss were recorded. Saliva was obtained and Ki-67 was estimated with a commercially available ELISA kit. Results The periodontitis group had significantly higher levels of Ki-67 than the healthy group. Overall, except with plaque index, there were significant positive weak correlations between Ki-67 and modified gingival index, bleeding on probing, probing pocket depth, and significant positive strong correlation with clinical attachment loss. Receiver operating characteristic analysis of the diagnostic accuracy of Ki-67 in periodontitis showed that the area under the curve was statistically significant, and the cutoff value was > 787.67 pg/ml. Conclusion Ki-67 can be detected in saliva and has a role in periodontitis. This implies that saliva can be a non-invasive source for Ki-67 as a potential biomarker in periodontitis.