Mass Spectrometric Analyses of Peptides and Proteins in Human Gingival Crevicular Fluid

Abstract

Gingival crevicular fluid (GCF) is a pathophysiological fluid that flows into the oral cavity. Human GCF was collected using sterile glass microcapillary tubes from inflamed periodontal sites in patients who had a history of periodontal disease and were in the maintenance phase of treatment. Samples from individual sites were analyzed using MS techniques both before and following HPLC. GCF samples were also pooled and subjected to SDS-PAGE, in-gel digestion and MS analyses using both MALDI-TOF/TOF MS and nanoLC-ESI-MS/MS. MS spectra were used to search human protein sequence databases for protein identification. With these approaches, 33 peptides and 66 proteins were positively identified in human GCF. All of the peptides discovered in this study are reported in GCF here for the first time. Forty-three of the identified proteins, such as actin and the actin binding proteins profilin, cofilin and gelsolin, have not been reported in GCF before.

C-Terminal Domain Residues Important for Secretion and Attachment of RgpB in Porphyromonas gingivalis

“…Gels were stained using Coomassie blue R250 (Bio-Rad, Hercules, CA) or SYPRO Ruby (Molecular Protein identification. Proteins were identified in SDS-PAGE gels using in-gel digestion and mass spectrometry (MS) using matrix-assisted laser desorption ionization-two-stage time of flight mass spectrometry (MALDI-TOF/TOF MS) as described previously (18). MS spectra were used to search a P. gingivalis protein sequence database for protein identification, as described previously (40).…”

Human infant saliva peptidome is modified with age and diet transition

“…Therefore, in the past few years, peptides have been studied by MS in whole saliva [1,2], parotid saliva [3], glandular secretory granules [4] and gingival crevicular fluid [5,6]. Salivary peptide profiles may be the mirror of important oral pre-and post-secretory biochemical reactions such as the proteolysis/proteolysis inhibition balance [7] or other posttranslational modifications.…”

Gingival Crevicular Fluid as a Novel Potential Source of Biomarkers Distinguishes Pubertal from Post-Pubertal Subjects

“…The total amounts of proteins and peptides that have been identified in GCF in periodontal healthy subjects are reported to be in a range from 88 [29] to 327 [30] that depends on the different sample sizes and methods employed in different studies. Many studies have applied gel-based or gel-free mass spectrometric analysis of GCF proteomic profiling for the detection of novel biomarkers of periodontitis and have successfully discovered several protein biomarkers that have never been investigated before [20,21,31], and some have confirmed the significance of biomarkers that had been proposed in previous studies via conventional research methods [30]. …”

“…However, the analysis of GCF is still challenging for limited volumes (0.2–1.5 μL per site) [21] due to the lack of a standardized protocol for collection and processing [23]. There are still some limitations in our work.…”

“…Ngo et al conducted MALDI-TOF/MS analysis of GCF and screened for several mass peaks to predict the sites with attachment loss [20]. Luan et al used MALDI-MS and LC-MALDI-MS/MS of undigested GCF to determine the peptide composition of GCF and successfully detected 33 peptides [21]. However, no studies have employed GCF proteomic analysis to identify novel biomarkers for pubertal growth spurt so far.…”