Figure 5.

Selective reaction monitoring (SRM) mass spectrometry analysis. The peptide VISSIEQKTMAAPSVK at the junction site of the BF969911.1 chimera was confirmed by SRM analysis using a stable isotope labeled standard. Briefly, a peptide of the same amino acid sequence was synthesized with a heavy lysine residue, which was then spiked into the digested human prostate cancer lysate. The mixture was fractionated by high pH reversed phase liquid chromatography and the fractions analyzed by SRM mass spectrometry. On the basis of the concentration of the labeled standard, the chimera was estimated to be present at a concentration of ∼30 fmol/mL. A signal-to-noise ratio was calculated as root-mean-square (RMS).