Detection of Ca2+-Binding S100 Proteins in Human Saliva by HPLC-ESI-MS
High-performance liquid chromatography (HPLC) coupled with electrospray ionization (ESI) mass spectrometry (MS) is a relevant technique for the detection and relative quantitation of naturally occurring peptides and proteins. The peptide/protein mass is determined by deconvolution of the ESI-MS spectrum, and the resolution can be better than 1:10,000 with the instruments currently available. Accurate mass measurement, coupled with sufficient resolution, makes it possible to greatly restrict the enormous number of possible molecular formulas that might be represented by a specific molecular mass.
As soon as the protein mass has been unequivocally attributed to a specific structure by means of different enzymatic and chemical treatments, the m/z values detected in the ESI spectrum can be utilized to reveal the protein and to perform its relative quantitation, by the extracted ion current (XIC) procedure, in an unlimited number of samples. This chapter describes the HPLC-ESI-MS experimental conditions which allow detecting and quantifying—in human saliva—different S100 proteins and their isoforms.
Key wordsHuman saliva HPLC-ESI-MS Label-free quantification XIC procedure S100 proteins EF-hand Calcium Post-translational modifications
We acknowledge the financial support of the Università di Cagliari, Università Cattolica, MIUR, the Italian National Research Council (CNR) and Regione Sardegna. We thank them for their programs on the promotion of scientific research and diffusion.
- 1.Covey T (1996) Analytical characteristics of the electrospray ionization process. Chapter 2. In: Snyder AP (ed) Biochemical and biotechnological applications of electrospray ionization mass spectrometry. ACS Symposium Series, vol 619, Oxford University Press, Oxford, pp 21–59Google Scholar