pFind Studio: a computational solution for mass spectrometry-based proteomics

Introduction

In relative protein abundance determination from peptide intensities recorded in full mass scans, a major complication that affects quantitation accuracy is signal interference from co-eluting ions of similar m/z values. Here we present pQuant, a quantitation software tool that solves this problem. pQuant detects interference signals, identifies for each peptide a pair of least interfered isotopic chromatograms — one for the light and one for the heavy isotope-labeled peptide. Based on these isotopic pairs, pQuant calculates the relative heavy/light peptide ratios along with their 99.75% confidence intervals (CIs). From the peptides ratios and their CIs, pQuant estimates the protein ratios and associated CIs by kernel density estimation. We tested pQuant, Census and MaxQuant on datasets obtained from mixtures (at varying mixing ratios from 10:1 to 1:10) of light- and heavy-SILAC labeled HeLa cells or 14N- and 15N-labeled E. coli cells. pQuant quantitated more peptides with better accuracy than Census and MaxQuant in all 14 datasets. On the SILAC datasets, the non-quantified “NaN” (Not a Number) ratios generated by Census, MaxQuant and pQuant accounted for 2.5–10.7%, 1.8–2.7% and 0.01–0.5% of all ratios, respectively. On the 14N/15N datasets, which cannot be quantified by MaxQuant, Census and pQuant produced 0.9–10.0% and 0.3–2.9% “NaN” ratios. Excluding these “NaN” results, the standard deviations of the numerical ratios calculated by Census or MaxQuant are 30–100% larger than those by pQuant. These results show that pQuant outperforms Census and MaxQuant in SILAC and 15N-based quantitation.




Supplemental Files

Our paper pQuant Improves Quantitation by Keeping out Interfering Signals and Evaluates the Accuracy of Calculated Ratios should be online. The identified/quantified peptides and proteins along with their calculated ratios and Raw files in this paper are available at ftp://PASS00466:ZG9287m@ftp.peptideatlas.org/.

pFind, pQuant and other software tools are integrated together in pFindStudio 3.0. We have released a private beta version.

Also, you can download pQuant.