title: Comparison of diverse affinity based high-abundance protein depletion strategies for improved bio-marker discovery in oral fluids.
abstract: Oral fluids (OF) have been suggested as a source of biomarkers for oral and systemic health, but as with other bio-fluids, the presence of high-abundance proteins interferes with the detection of lower-abundance biomarkers. Here, we compared the performance of four depletion treatments: triple depletion (TD) of amylases, albumins and immunoglobulin G; multiple depletion (MD) of amylases and a panel of 20 proteins, a combination of the two (EMD) and combinatorial peptide ligand library based depletion termed CPLL. TD, MD, EMD and CPLL removed 76%, 83%, 85% and 94% of total proteins, respectively, coupled with increased low abundance protein detection and narrowed dynamic range. 2-DE revealed that all depletion pretreatments successfully clarified areas hampered by high-abundance proteins; however, EMD and CPLL exposed the highest number of proteins. Quantitative MS of EMD samples relative to none treated samples indicated that most of downregulated proteins (>90%) were EMD target proteins. In conclusion, a multiple step EMD and CPLL depletion approaches bring about the highest number of protein detection ability and the best hampered-area clearance. As CPLL requires at least 10 fold more protein starting material, we suggest EMD pretreatment as a new detection tool in instances of low protein starting material.
authors.affiliation: Institute of Dental Sciences, The Hebrew University, Israel.
authors.forename: Guy
authors.initials: G
authors.lastname: Krief
authors.forename: Omer
authors.initials: O
authors.lastname: Deutsch
authors.forename: Batia
authors.initials: B
authors.lastname: Zaks
authors.forename: David T
authors.initials: DT
authors.lastname: Wong
authors.forename: Doron J
authors.initials: DJ
authors.lastname: Aframian
authors.forename: Aaron
authors.initials: A
authors.lastname: Palmon
mesh_terms: D000328:Adult; D015415:Biomarkers / Q000032:analysis*; D001826:Body Fluids; D020650:Combinatorial Chemistry Techniques; D015180:Electrophoresis, Gel, Two-Dimensional; D006801:Humans; D008297:Male; D011506:Proteins / Q000302:isolation & purification*; D040901:Proteomics / Q000379:methods*; D012463:Saliva / Q000737:chemistry*
chemical_list: D015415:Biomarkers; D011506:Proteins