Yuzhe Nie (Presenter)
University of South Carolina
Bio: Yuzhe Nie received his Bachelor"s degree in Biology in 2003, M.S. in Ornamental Plant and Horticulture in 2006 and D.Sc. in Developmental Biology in 2010 from Northeast Forestry University, China. He studied proteomics at Functional Proteomics Key Lab of Guangdong Province, China for protein and peptides assay by mass spectra technology in 2008; awarded Finland CIMO's scholarships to do a 6 month joint research on environmental bioremediation at University of Helsinki in 2009; After graduating, he worked as an assistant researcher focusing on disease-detection protein marker screening by 2D-DIGE and mass spectra technology at Northeast Forestry University. Currently, he is a visiting post doctoral fellow in Chemistry and Biochemistry Department at University of South Carolina mainly working on mass spectrometric analysis of peptides, glycosylation profiling, post-translational modificat
Authorship: Yuzhe Nie(1), L. Andrew Lee(2), Qian Wang(1)
(1) University of South Carolina, (2) IMCS, LLC
| Short Abstract For clinical proteomics, targeted peptides/proteins extraction and desalting efficiency are essential to obtaining high quality mass spectra data. In this study, we report a novel peptide purification method using IMCStipsTM, a disposable pipette extraction tip with sorbent loosely contained inside a pipette tip. Using human serum albumin as a model protein, the trypsin digested samples were readily purified using IMCStips™ packaged with reverse phase resin. The purification improved MS peak intensities, S/N ratios and sequence coverage significantly in comparison to other commercially available tips. Similar results have been obtained with other proteins, like human prealbumin, ¦Á-acid-glycoprotein and transferrin. Since this method can be used in an automated online format, we envision it to be applied in the clinical proteomics studies, especially for low abundance peptides. |
Long Abstract
For clinical proteomics, it is critical to enrich peptide samples in a high-throughput fashion. Since we often deal with disease relevant low abundant proteomes, the extraction and desalting efficiency of targeted peptides/proteins play an essential role in getting high quality mass spectra data. Traditional solid-phase extraction (SPE) pipette extraction tips were usually used in this task, but at larger scale, the proportional increased in solvent volumes and liquid resistant force would limit the application on large scale high-throughput proteomics samples.
In this study, we report a novel peptide purification method using IMCStips, which based on sorbent loosely contained inside pipette tips to increase dispersive mixing between resin and sample within the tips. The dispersive nature of the resin promotes sample homogenization by forcing the resin to mix with the solution. This reduces channeling effects often seen with conventional columns, and results in improved recovery as samples are in contact with more surface area during the dispersive mixing stage. Furthermore, bidirectional flow means that solvent can be introduced and removed from the tip more rapidly, speeding the extraction process. Lastly, due to the dispersion process, smaller bed volume is needed. This allows the user to reduce the amount of solvent used per extraction. The dispersive pipette extration tips have been proven with the ¡°QuEChERS Tips¡± and ¡°DPX-WAX¡± which can purify pesticides and organic acids effectively and quickly[1,2].
Using purified human serum albumin (purchased from Athens Research & Technology) as a model protein, the trypsin-digested samples were readily purified using reverse phase resin. To test the purification effect, resultant peptide samples were assayed by MALDI TOF. Comparing the MS results before and after desalting, trypsin-digested human albumin's MS peak intensity improve 3.5 folds, S/N improved 8.3 fold and sequence coverage improved from 50% to 68%, the result is better than ZipTip® C18 Pipette Tips which was now widely used on peptide desalting[3]. Similar results have been obtained with other proteins, prealbumin, ¦Á-acid-glycoprotein and transferrin (Athens Research & Technology). After desalting, their MS peak intensities improved 4.2, 4.2 and 11.0 fold, S/N improve 3.4, 2.1 and 2.4 fold, sequence coverage improve from 33% to 48%, 46% to 47%, 30% to 34% respectively. By using multichannel pipette, the above trypsin-digested protein"s purification could be finished in several minutes, also it can be expanded to complete online automation for higher throughput labs. We envision it be applied in the clinical proteomics studies using more disease related peptides, especially for weak signal peptides.
References & Acknowledgements:
[1] Pay¨¢, Paula, et al. "Analysis of pesticide residues using the Quick Easy Cheap Effective Rugged and Safe (QuEChERS) pesticide multiresidue method in combination with gas and liquid chromatography and tandem mass spectrometric detection." Analytical and bioanalytical chemistry 389.6 (2007): 1697-1714.
[2] Guan, Hongxia, William E. Brewer, and Stephen L. Morgan. "New approach to multiresidue pesticide determination in foods with high fat content using disposable pipette extraction (DPX) and gas chromatography− mass spectrometry (GC-MS)." Journal of agricultural and food chemistry 57.22 (2009): 10531-10538.
[3] Aresta, Antonella, et al. "Impact of sample preparation in peptide/protein profiling in human serum by MALDI-TOF mass spectrometry." Journal of pharmaceutical and biomedical analysis 46.1 (2008): 157-164.
| Description | Y/N | Source |
| Grants | yes | IMCS, LLC |
| Salary | no | |
| Board Member | no | |
| Stock | no | |
| Expenses | no |
IP Royalty: no
| Planning to mention or discuss specific products or technology of the company(ies) listed above: | yes |