Enhanced Analysis of Metastatic Prostate Cancer Using Stable Isotopes and High Mass Accuracy Instrumentation

The primary goal of proteomics is to gain a better understanding of biological function at the protein
expression level. As the field matures, numerous technologies are being developed to aid in the
identification, quantification and characterization of protein expression and post-translational modifications on a near-global scale. Stable isotope labeling by amino acids in cell culture is one such technique
that has shown broad biological applications. While we have recently shown the application of this
technology to a model of metastatic prostate cancer, we now report a substantial improvement in
quantitative analysis using a linear ion-trap Fourier transform ion cyclotron resonance mass spectrometer (LTQ FT) and novel quantification software. This resulted in the quantification of nearly 1400
proteins, a greater than 3-fold increase in comparison to our earlier study. This dramatic increase in
proteome coverage can be attributed to (1) use of a double-labeling strategy, (2) greater sensitivity,
speed and mass accuracy provided by the LTQ FT mass spectrometer, and (3) more robust quantification
software. Finally, by using a concatenated target/decoy protein database for our peptide searches, we
now report these data in the context of an estimated false-positive rate of one percent.
Keywords: mass spectrometry • SILAC • prostate cancer • quantitative proteomics