MS Bioworks is proud to announce our first Q Exactive instrument is on-line and available for client projects.
What is the Q Exactive?
The Thermo Scientific Q Exactive is a hybrid quadrupole-Orbitrap mass spectrometer combining a high-performance quadrupole mass filter with the industry leading high resolution sensitive and selective Orbitrap mass analyzer.
What is the Q Exactive advantage?
Speed
Q Exactive allows for 10 MS/MS spectra per second to be acquired, resulting in an unprecedented number of protein identifications. The data below were generated from the analysis of 1µg of a K562 cell lysate. Each sample was analyzed in triplicate. The K562 tryptic digest was analyzed using 0.5, 1, 2, 3 and 4hr reverse phase gradients. Protein identification was performed using MaxQuant 1.2.2.5 with the Andromeda search engine. More information on the experimental methods can be found at the bottom of the page.
Analysis Time (hrs) | 0.5 | 1 | 2 | 3 | 4 | |||||
---|---|---|---|---|---|---|---|---|---|---|
average | % CV | average | % CV | average | % CV | average | % CV | average | % CV | |
Number of identified Proteins (with two unique peptides per protein) |
970.5 | 3.0 | 1960 | 0 | 2332 | 1.3 | 2812.5 | 2.1 | 3028.5 | 2.2 |
The data above shows the number of proteins identified as a function of analysis time.
Analysis Time (hrs) | 0.5 | 1 | 2 | 3 | 4 | |||||
---|---|---|---|---|---|---|---|---|---|---|
average | % CV | average | % CV | average | % CV | average | % CV | average | % CV | |
Unique Peptides | 5458 | 2.9 | 13529 | 0.4 | 18101.5 | 0.5 | 20563 | 3.4 | 23496 | 0.3 |
The data above shows the number of unique peptides identified as a function of analysis time.
Analysis Time (hrs) | 0.5 | 1 | 2 | 3 | 4 | |||||
---|---|---|---|---|---|---|---|---|---|---|
average | % CV | average | % CV | average | % CV | average | % CV | average | % CV | |
PSMs | 6865.5 | 5.7 | 17392 | 0.8 | 23648 | 0.5 | 26684 | 6.1 | 30725.5 | 0.3 |
The data above shows the number of Peptides Spectra Matching (PSM) identified as a function of analysis time. A PSM is any spectrum matching to a peptide and includes repeat observations and PTM-modified peptides.
Accuracy
The Q Exactive acquires the precursor and product ion masses at high resolution. The data can be used to accurately and efficiently assign the site of a modification. In the example below a phosphorylation site is confidently localized based on the MS/MS spectra.
(K)YNLDAsEEEDSNK(K) from Zinc finger Ran-binding domain-containing protein 2
To find out how the Q Exactive can impact your proteomics research contact MS Bioworks today and speak with a scientist.
Email: info@MSBioworks.com or call 734-929-5083
Experimental Details
Mass Spectrometry
The tryptic digest was analyzed by nano LC/MS/MS with a Waters NanoAcquity HPLC system interfaced to a ThermoFisher Q Exactive mass spectrometer. Peptides were loaded on a trapping column and eluted over a 75µm analytical column at 350nL/min; both columns were packed with Jupiter Proteo resin (Phenomenex). Gradients of 0.5, 1, 2, 3 and 4h were employed in analytical triplicate. The mass spectrometer was operated in data-dependent mode, with MS performed at 70,000 FWHM resolution and MS/MS performed at 17,500 FWHM resolution. The fifteen most abundant ions were selected for MS/MS.
Data Processing
Data were searched using a local copy of MaxQuant 1.2.2.5 with the Andromeda search engine using the following parameters:
Enzyme: Trypsin
Database: SwissProt Human (appended with common contaminants, reversed and concatenated)
Fixed modification: Carbamidomethyl (C)
Variable modifications: Oxidation (M), Acetyl (N-term), Pyro-Glu (N-term Q), Deamidation (NQ)
Mass values: Monoisotopic
Peptide Mass Tolerance: 6 ppm
Fragment Mass Tolerance: 20 ppm
Max Missed Cleavages: 2
Peptide FDR: 1%
Protein FDR: 1%
Min Unique Peptides: 2