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AQUA™ Depletion plasma facts
Figure 2.Plasma Protein Depletion Workflow
High-Abundance Protein Depletion
An AQUA™ Peptide stock systems was ready by dissolving isotopically labeling versions that the 2 target peptides (FQNALLVR* and also GASQAGAPQGR*, respectively) in 0.1% TFA to a final concentration of 62.5 fmol/mL. Every digest sample (whole and also depleted plasma) was liquified in 20 mL that the AQUA™ Peptide share solution. Samples to be analyzed making use of an Agilent capillary 1100 HPLC coupled to a Thermo Finnigan LTQ linear ion trap mass spectrometer. Utilizing an LC-SRM method, the absolute amount of each peptide (and matching protein) was determined.
Figure 3. Overview the the Protein-AQUA™ Method. This method was arisen by Dr. Steve Gygi and also colleagues at Harvard medical School
Select an optimal tryptic peptide and also stable isotope amino acid from the succession of her protein of interest.
Order artificial AQUA™ Peptide from.
Optimize LC-MS/MS separation protocol because that quantitation.
Extract protein from biological samples and add known quantity of AQUA™ peptide.
Analyze by LC-MS/MS or MALDI to quantitate protein of interest.
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Anderson NL, Anderson NG. 2002. The human Plasma Proteome. Mol cell Proteomics. 1(11):845-867. Http://dx.doi.org/10.1074/mcp.r200007-mcp200
Gerber SA, rush J, Stemman O, Kirschner MW, Gygi SP. 2003. Absolute quantification that proteins and phosphoproteins from cell lysates through tandem MS. Proceedings the the nationwide Academy that Sciences. 100(12):6940-6945. Http://dx.doi.org/10.1073/pnas.0832254100