Here, we investigate how delays in plasma processing affect peptide-centric “bottom-up” proteomics, using multiple reaction monitoring mass spectrometry (MRM-MS). We used validated Tier 2-level assays for proteotypic peptide surrogates of 270 human proteins to analyze plasma generated from whole blood that has been kept at room temperature from 0-40 hours before plasma generation. Moreover, we evaluated whether different plasma thawing conditions had an impact on the MRM-based plasma protein measurements.