Methods & Services
Quantitative Proteomics
The Core Facility for Mass Spectrometry and Proteomics offers several quantification strategies:
Label free using MaxQuant software (http://www.ncbi.nlm.nih.gov/pubmed/20171123) - see Figure 1A
Chemical labeling at the peptide level using Reductive Dimethylation (DML) (http://www.ncbi.nlm.nih.gov/pubmed/19300442) - see Figure 1B
Metabolic labeling using SILAC (http://www.ncbi.nlm.nih.gov/pubmed/17406521) - see Figure 1C
Targeted proteomics using selective/multiple reaction monitoring (SRM/MRM) (http://www.ncbi.nlm.nih.gov/pubmed/18854821) or parallel reaction monitoring (PRM) (http://www.ncbi.nlm.nih.gov/pubmed/22865924) - see Figure 2
Absolute quantification using AQUA peptides (http://www.ncbi.nlm.nih.gov/pubmed/15722223)
Unbiased proteomic studies can be performed using our high resolution Orbitrap mass spectrometers to provide accurate quantification data. For all targeted proteomics approaches, a triple quadrupole mass spectrometer is recommended, as MRM (multiple reaction monitoring) is inherently more sensitive compared to regular MS/MS.
Quantification strategies
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B
C
Figure 1 - Quantification strategies available in the Core Facility
Targeted proteomics using MRM
We also perform targeted analysis, namely MRM (Qtrap 5500), to determine protein complex stoichiometries. The working principle of MRM is shown in Figure 2.
Benchmarking our quantitative MS performance - Succesful participation in ABRF sPRG study 2014
In 2014 we participated in a cross-laboratory study trying to access their quantitative capacities. Therefore a standardized sample containing isotope labeled peptides in a complex HEK293 background was distributed to all participants. As one can observe from the results, our core facility performed very well, especially when using MaxQuant as data analysis software.
