Is Subtractive Affinity Depletion of Abundant Serum Proteins Useful and Reproducible?
Proteomics: Sample Preparation and Methodologies I Sample
enrichment by abundant protein depletion was found to expand the
effective dynamic range of LC/MSE analysis by up to two orders of
magnitude protein concentration.Both the Top6 and Top20 depletion
workflows appear robust, and sufficient quantitative capability is
retained to permit depleted serum to be used for biomarker
identification. Samples processed using Top20 depletion produced
significantly more protein identifications than the comparable Top6
depletion strategy. The high sequence coverage and reproducible peptide
intensities obtained from LC/MSE analysis permitted confident
calculation of protein depletion efficiencies. In some cases,
biologically relevant protein fragments could be enriched and detected
following the targeted removal of the intact form(s) of the protein.
Overall, targeted affinity depletion of high abundance serum proteins
appeared to be an effective sample preparation strategy prior to LC/MSE
analysis by the Waters Protein Expression System.