Large-scale multiplexed quantitative discovery proteomics enabled by the use of an (18)O-labeled "universal"...

Area of Science:

• Proteomics
• Quantitative Mass Spectrometry
• Biomarker Discovery

Background:

• Accurate protein quantification across numerous samples is crucial for proteomics discovery.
• Existing methods face challenges with large sample sets and instrument variability.

Purpose of the Study:

• To introduce and validate a novel quantitative proteomics strategy using a stable isotope-labeled universal reference sample.
• To assess the performance of this method for large-scale sample analysis, including clinical samples.

Main Methods:

• Incorporation of an oxygen-18 ((18)O)-labeled universal reference sample as internal standards.
• Quantification based on (16)O/(18)O isotopic peptide pair abundance ratios.
• Simultaneous application of label-free quantitation (dual-quantitation) for comparison.
• Analysis of 18 plasma samples from severe burn patients using LC-MS/MS after immunoaffinity depletion and cysteinyl-peptide enrichment.

Main Results:

• A total of 312 plasma proteins were identified and quantified.
• The (18)O reference-labeling approach demonstrated significantly better quantitative precision than label-free quantitation.
• Strong correlation was observed between (18)O labeling and label-free quantitation results, highlighting complementary strengths.

Conclusions:

• The (18)O-labeled universal reference sample strategy offers a simple and accurate method for large-scale quantitative proteomics.
• This approach enhances precision, especially when instrument robustness is a concern for label-free methods.
• The method is valuable for systems biology studies aiming to discover subtle protein abundance changes.