Discovery Proteomics and Absolute Protein Quantification Can Be Performed Simultaneously on an Orbitrap-Based Mass...

Area of Science:

• Proteomics
• Analytical Chemistry
• Biochemistry

Background:

• Mass spectrometry (MS) is crucial for protein quantification.
• Isotope dilution MS with peptide fragments is a standard quantification technique.
• Parallel reaction monitoring (PRM) is often preferred for Orbitrap instruments due to assumed sensitivity gains.

Purpose of the Study:

• To compare data-dependent acquisition (DDA) and PRM workflows for quantifying vitellogenins (VTGs) in striped bass.
• To assess VTG proportions as a measure of egg quality and environmental impact.
• To determine if DDA can provide both discovery and absolute quantification data.

Main Methods:

• Comparative analysis of DDA and PRM-based workflows.
• Quantification of vitellogenins (VTG) Aa, Ab, and C in striped bass (Morone saxatilis).
• Statistical comparison using single-factor analysis of variance (ANOVA).

Main Results:

• No statistically significant difference was found between MS1-based (DDA) and MS2-based (PRM) VTG quantification.
• DDA workflows successfully provided both protein discovery and absolute quantification data within a single experiment.
• VTG proportions are a potential indicator of fish stock health and environmental changes.

Conclusions:

• DDA is a viable and efficient method for absolute protein quantification, comparable to PRM.
• DDA offers a dual benefit of discovery and quantification, streamlining proteomic experiments.
• VTG quantification via MS can inform ecological and developmental studies in fish populations.