Abstract
BackgroundHigh abundance protein depletion is a major challenge in the study of serum/plasma proteomics. Prior to this study, most commercially available kits for depletion of highly abundant proteins had only been tested and evaluated in adult serum/plasma, while the depletion efficiency on umbilical cord serum/plasma had not been clarified. Structural differences between some adult and fetal proteins (such as albumin) make it likely that depletion approaches for adult and umbilical cord serum/plasma will be variable. Therefore, the primary purposes of the present study are to investigate the efficiencies of several commonly-used commercial kits during high abundance protein depletion from umbilical cord serum and to determine which kit yields the most effective and reproducible results for further proteomics research on umbilical cord serum.ResultsThe immunoaffinity based kits (PROTIA-Sigma and 5185-Agilent) displayed higher depletion efficiency than the immobilized dye based kit (PROTBA-Sigma) in umbilical cord serum samples. Both the PROTIA-Sigma and 5185-Agilent kit maintained high depletion efficiency when used three consecutive times. Depletion by the PROTIA-Sigma Kit improved 2DE gel quality by reducing smeared bands produced by the presence of high abundance proteins and increasing the intensity of other protein spots. During image analysis using the identical detection parameters, 411 ± 18 spots were detected in crude serum gels, while 757 ± 43 spots were detected in depleted serum gels. Eight spots unique to depleted serum gels were identified by MALDI- TOF/TOF MS, seven of which were low abundance proteins.ConclusionsThe immunoaffinity based kits exceeded the immobilized dye based kit in high abundance protein depletion of umbilical cord serum samples and dramatically improved 2DE gel quality for detection of trace biomarkers.
Highlights
High abundance protein depletion is a major challenge in the study of serum/plasma proteomics
To set up a high abundance protein depletion method suitable for umbilical cord serum, we evaluated the three most commonly used depletion kits, testing for depletion efficiency, reproducibility, and the yield of depleted serum
The Blue Albumin and IgG Depletion kit (PROTBA, Sigma-Aldrich, Saint Louis, MO, USA) is an immobilized dye based depletion kit which is highly efficient in depleting albumin and IgG from adult plasma/serum at a low cost
Summary
High abundance protein depletion is a major challenge in the study of serum/plasma proteomics. Most commercially available kits for depletion of highly abundant proteins had only been tested and evaluated in adult serum/plasma, while the depletion efficiency on umbilical cord serum/plasma had not been clarified. Structural differences between some adult and fetal proteins (such as albumin) make it likely that depletion approaches for adult and umbilical cord serum/plasma will be variable. For plasma/serum based proteomics research, the depletion of high abundance proteins is of great importance in sample preparation. Several depletion kits have been developed to remove highly abundant proteins in human plasma/serum [6,7,8,9] for use in further proteomics research. Most of the depletion approaches have only been tested and evaluated in adult serum or plasma, and little data about their usage and efficiency in fetal or umbilical cord serum/ plasma depletion was available
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
