Proceedings book download - 5Z.com

Table 1. The quantity of identified proteins and peptides for each group of samplesExperimental groupsNumber of identifiedproteinsNumber of identifiedpeptidesHealthy donors 290 656Ovarian cancer 178 462Colorectal cancer 137 354Syphilis 158 452Daltonics, Germany). The best generated classification models demonstrated the followingvalues of sensitivity and specificity for the detection of studied diseases: ovarian cancer –100% and 100%; colorectal cancer – 100% and 100%; syphilis – 92% and 100% (forsensitivity and specificity, respectively).Serum samples of all experimental groups were subjected to consecutive separation byWCX and SAX chromatography followed by RP-LC-MS/MS analyses using the Agilent1200 HPLC-Chip/MS Interface coupled with the Agilent 6520 Accurate-Mass Q-TOF. Asa result of MS/MS identification of peptides that was performed by PhenyxServer (GenevaBioinformatics S.A.) using databank Uniprot-SwissProt 1087 unique peptides representing487 proteins were reliably identified (see Table 1) (validation of peptide identifications wasmade by searching MS/MS data against the reversed version of Uniprot-SwissProt).LC-MS/MS analysis of serum samples revealed significant discrepancy betweenidentified peptides and proteins in various experimental groups that contradict visualsimilarity of MALDI-TOF MS profiles between the same groups. Resolution of linearmode of MALDI-TOF mass spectrometers is too low for detail analyses of such complexpeptide mixture as serum is, while a reflector mode is difficult to use because of its lowersensitivity. From the obtained results it is also obvious that it is impossible to correlate thedata of LC-MS/MS identifications with MS peaks of potential biomarkers revealed fromMALDI-TOF MS profiling of serum samples. We believe that high-throughputcomparative MALDI-TOF MS profiling of serum samples can be used for preliminaryestimation of differences between peptide profiles of tested groups, with followingdetection of peptide biomarkers using label-free quantitative LC-MS approach and theirfurther identification by LC-MS/MS.AcknowledgmentsThis work is supported by Russian Foundation for Basic Research (09-04-12085).References1. Anderson, N.L., Anderson, N.G. Mol. Cell. Proteomics 1, 845-867 (2002).2. Mehta, A.I., Ross, S., Lowenthal, M.S., Fusaro, V., Fishman, D.A., Petricoin, E.F., 3 rd , Liotta, L.A.Dis. Markers 19, 1-10 (2003).3. Geho, D.H., Liotta, L.A., Petricoin, E.F., Zhao, W., Araujo, R.P. Curr. Opin. Chem. Biol. 10, 50-55(2006).277