Fig. 2.

Comparison of MS1 (pulsed SILAC) and MS3 (pulsed SILAC-TMT) based quantification. A, The fraction of MS3 spectra as a function of the detected ratio compression (from measuring the residual intensities in outermost TMT channels) illustrates that ratio distortion was still present, but that >80% of all fitted and filtered spectra showed less than 10% residual intensities. B, Correlation analysis of log transformed labeling rates showed good agreement between the MS1 and MS3 based quantification approaches (R: Pearson's correlation coefficient). C, Labeling characteristics measured for the protein STAT3 either using the MS1 or MS3 strategy yielded consistent data. D, Fractional labeling determined for the protein STAT6 in which MS1 data points were missing (one SILAC isotope pair signal missing for 1, 3, 6, and 48 h time points and no data for the 10 and 24 h time points) led to substantial differences in curve fits between MS1 and MS3 data. E, Distributions of coefficients of determination (R²) of curve fits display consistently higher values for the MS3 compared with the MS1 approach (dotted lines: medians). F, Comparison of the number of proteins with determined turnover parameters shows a higher number for the MS3 strategy.