poor coverage of target transcription factor proteins in nist database supplementary figure 2 . in vitro - synthesized proteins are enriched full - length proteins supplementary figure 3 . histogram of dot - products for quality score 1 and 2 peptides supplementary figure 5 . peptide ms / ms spectrum counts are a poor predictor of targeted peptide signal intensity using selected reaction monitoring - mass spectrometry supplementary figure 7 . comparison of our empirical rankings with peptide rank predictions from the espp redictor algorithm supplementary note . skyline tutorial on protein absolute quantification using a heavy isotope internal standard and an external calibration curve
we report a method for high - throughput , cost - efficient empirical discovery of optimal proteotypic peptides and fragment ions for targeted proteomics applications using in vitro - synthesized proteins . 
 we demonstrate the approach using human transcription factors  which are typically difficult , low - abundance  targets with an overall success rate of 98% . 
 we show further that targeted proteomic assays developed using our approach facilitate robust in vivo quantification of human transcription factors .