Use of ProteinChip™ array surface enhanced laser desorption/ionization time-of-flight mass spectrometry (SELDI-TOF MS) to identify thymosin β-4, a differentially secreted protein from lymphoblastoid cell lines

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Abstract
The identification of proteins differentially expressed between cancer and normal cells is vital for the development of cancer diagnostics, therapeutics and vaccines. Using a ProteinChip Biomarker System (Ciphergen Biosystems, Fremont, CA) which combines ProteinChip™ technology with time-of-flight mass spectrometry, we have developed a simple method to screen and identify differentially secreted proteins from tumor cell lines. Mass spectra of the range of proteins secreted from normal B-cells were generated along with those secreted from Epstein-Barr virus transformed B-cells. A mass peak at m/z = 4972. 1 that was highly over-represented in the transformed B-cell line was chosen for identification and purified by reversed phase chromatography with concomitant monitoring of fractions by SELDI-TOF MS. The resulting purified protein was digested with trypsin and the peptide masses derived from the SELDI-TOF spectrum were used to search the public databases for protein identification. Fragment matching of the resulting peptides identified the protein as thymosin β-4. Using LC-electrospray ionization MS/MS, the identity of this protein was confirmed. Thymosin β-4 is a known marker in LCLs establishing the utility of this method to discover and identify proteins differentially expressed between cancers and their matched normal counterparts.