Comparative proteomic analysis of saliva and salivary stones in sialolithiasis

The proteins associated with poorly described mechanism of sialolithiasis pathogenesis were previously described, but to increase credibility of these results and to discover new biomarkers of this disease it would be beneficial to verify the validity of optimized protocols during performing the quantitative analysis to establish the most reasonable reference sample. Previously established protocols were used to perform optimal protein extraction and digestion in saliva and salivary stone samples. Based on the DDA spectra the well-developed spectra library was created and then the DIA spectra were used to conduct relative quantitative proteomic analysis of saliva and sialoliths. The optimized workflows allowed to quantify the proteins in saliva and salivary stone samples. After statistical analysis it was possible to compare protein profiles of different saliva samples and sialolith samples, depending on the chosen reference sample. This study verified the applicability of saliva as reference sample in quantitative proteomic analysis of sialoliths, but at the same time no differences between saliva from healthy donors and saliva from patients with sialolithiasis were detected. It was possible to identify high confident proteins associated with pathological calcification leading to sialoliths formation.