Over the last years, proteomic techniques have been demonstrated to be valuable tools for analysing the periodontal proteome in gingival crevicular fluid (GCF) samples contributing to identifying extensive and comprehensive protein profiles associated with periodontitis. However, the periodontal proteome has not yet been studied using sequential window acquisition of all theoretical mass spectra (SWATH-MS). Consequently, the objective of the present longitudinal research was to investigate the periodontal proteome in GCF in three clinical conditions (periodontal health, untreated periodontitis, and treated periodontitis), applying SWATH-MS to 1) analyse the periodontal proteome structure in the three clinical groups, 2) compare the differential protein expression between the three clinical groups, and 3) evaluate the diagnostic accuracy of the proteins quantified to identify new biomarkers with a high capacity to discriminate the three clinical conditions. A total of 84 subjects, 44 periodontally healthy and 40 with generalised untreated periodontitis in stages III-IV, were included, and GCF samples were collected. The diagnosis of periodontal status was conducted by two experienced dentists using clinical and radiographic criteria. The definition of periodontal status was established following the Classification of Periodontal Diseases and Conditions published in 2018. In the untreated periodontitis group, 38 underwent conventional non-surgical treatment and were evaluated clinically after two months (treated periodontitis group). In the revaluation, 25 individuals showed significant clinical improvement in probing pocket depth (PPD) at the selected sites, and GCF samples were collected from them.