IQUP identifies quantitatively unreliable spectra with machine learning for isobaric labeling-based proteomics.

Mass spectrometry‑based proteomics using isobaric labeling technology has become popular for proteomic quantitation. Existing approaches rely on the mechanism of target-decoy search and false discovery rate control to examine whether a peptide-spectrum match (PSM) is utilized for quantitation. However, some PSMs passing the examination may still exhibit high quantitation errors, which can deteriorate the overall quantitation accuracy. We present IQUP, a machine learning-based method to identify quantitatively unreliable PSMs, termed QUPs. PSMs were characterized by 16 spectral and distance-based features for machine learning. Independent test results reveal that the best-performing models for the three datasets achieve accuracies of 0.883-0.966, AUCs of 0.924-0.963, and MCCs of 0.596-0.691. Notably, the distributions of relative errors for QUPs and quantitatively reliable PSMs (QRPs) exhibit significant differences. By using only the predicted QRPs for peptide-level quantitation, the proportions of peptides with larger relative errors decrease significantly, with a range between 15.3 and 83.3% for the three datasets; in the meantime, the proportions of peptides with smaller relative errors increase by 3.1-25.5%. Our experimental results demonstrate that IQUP provides robust performance and strong generalizability across multiple datasets and has great potential in improving proteomic quantitation accuracy at PSM and peptide levels for isobaric labeling experiments.