Serum proteomics system and associated methods

A method for comparing multiple mass spectra from different biological samples, locating mass ions that are quantitatively different after using approaches to compensate for non-biological variability, and isolating and sequencing at least one peptide of interest thus allowing for identification of the peptide from a biological sample, comprising:

• fractionating each of a plurality of biological samples to form a plurality of elutions;
• obtaining a plurality of mass spectra from each of the plurality of elutions at a plurality of elution times;
• finding a molecular ion peak of interest that appears to be quantitatively different between biological samples;
• identifying a mass spectrum reference peak corresponding to an endogenous reference molecule that is substantially consistent between biological samples, the endogenous reference molecule having an elution time and a mass to charge ratio that are substantially similar to the peak of interest;
• compensating for non-biological variation for each biological sample across the plurality of elutions by normalizing the peak of interest to a mass spectrum peak of the endogenous reference molecule;
• and
• fractionating at least one of the biological samples containing the peptide associated with the peak of interest to at least partially isolate the peptide;
• obtaining mass spectra of the peptide;
• accelerating the peptide into a collision chamber at a plurality of discrete collision energies for a discrete period of time to form a plurality of peptide fragments for each of the plurality of discrete collision energies;
• obtaining a plurality of fragmentation mass spectra from the plurality of peptide fragments for each of the plurality of discrete collision energies;
• summing the plurality of fragmentation mass spectra from each of the plurality of discrete collision energies to form a plurality of discrete collision energy mass spectra, one discrete collision energy mass spectra from each discrete collision energy;
• summing the plurality of discrete collision energy mass spectra to form a final mass spectrum for the peptide fragments;
• and
• identifying a sequence of amino acids corresponding to the peptide from the final mass spectrum.