Targeted Proteomics Environment
description
Transcript of Targeted Proteomics Environment
Targeted ProteomicsEnvironment
Status of the Skyline open-source software project five years after its inception
Brendan MacLean
User Community After 4 Years 560 registered users 150 registered for this meeting 4 new papers in May
Platform Independent and Label-free Quantitation of Proteomic Data using MS1 Extracted Ion Chromatograms in Skyline – Mol. Cel. Prot.
Label-Free Quantitation of Protein Modifications by Pseudo-Selected Reaction Monitoring with Internal Reference Peptides – J. Prot. Res.
Using iRT, a Normalized Retention Time for More Targeted Measurement of Peptides - Proteomics
The Development of Selected Reaction Monitoring Methods for Targeted Proteomics via Empirical Refinement - Proteomics
25 abstracts at ASMS mention Skyline 75 citations of original paper (after 2 ½ years)
30 in 2012
User Community After 5 Years 1000+ registered users 250 registered for this meeting More papers and press
Targeted proteomics Nature Methods Method of the Year in 2012 MS1 Label-free Quantification Using Ion Intensity Chromatograms in
Skyline (Research and Clinical Applications) (book chapter) Design, Implementation, and Multi-Site Evaluation of a System
Suitability Protocol for the Quantitative Assessment of Instrument Performance in LC-MRM-MS - MCP
Viewing the Targeted Proteomics Horizon with Skyline – eProtein feature
37 abstracts at ASMS mention Skyline 200+ citations of original paper
Skyline Use
Dev 64 Release
Dev Release
1.3 Release1.2 Release1.1 ReleasePre-1.1 Release
1.3 64 ReleaseASMS
2010
ASMS2011
Skyline Instances Started Trailing 7 Days
Inst
ance
s
3500
3000
2500
1500
1000
500
0
2000
4000
4500
ASMS2012
5000
Panorama2013
Skyline Web Site Visits (past 3 months)
Learning More Au PS Workshop, Melbourne (February) US HUPO Workshop, Baltimore (March) Skyline User Group Meeting! here (now) SRM Course, Zurich (July) Proteomics Course, Cold Springs Harbor (July)
Targeted Quant. Course, Seattle
September 9-14
Instrument Vendor Partnerships
Panorama Partnership Program 2 – 4 labs Seeking leading edge targeted proteomics
infrastructure Local Panorama server installation Direct collaboration with Skyline/Panorama
team Full support for 1 year
Prior Knowledge and Consistency Powerful enough to be used cross-lab / cross
experiment More powerful run-to-run
Relative ion abundance Spectral and chromatogram libraries
Retention time iRT
Does ensuring comparable measurements require ID?
Haptoglobin Li
brar
y
D_1
02_R
EP
1
D_1
02_R
EP
2
D_1
02_R
EP
3
D_1
03_R
EP
1
D_1
03_R
EP
2
D_1
03_R
EP
3
D_1
08_R
EP
1
D_1
08_R
EP
2
D_1
08_R
EP
3
D_1
38_R
EP
1
D_1
38_R
EP
2
D_1
38_R
EP
3
D_1
54_R
EP
1
D_1
54_R
EP
2
D_1
54_R
EP
3
D_1
72_R
EP
1
D_1
72_R
EP
2
D_1
72_R
EP
3
D_1
96_R
EP
1
D_1
96_R
EP
2
D_1
96_R
EP
3
H_1
46_R
EP
1
H_1
46_R
EP
2
H_1
46_R
EP
3
H_1
47_R
EP
1
H_1
47_R
EP
2
H_1
47_R
EP
3
H_1
48_R
EP
1
H_1
48_R
EP
2
H_1
48_R
EP
3
H_1
59_R
EP
1
H_1
59_R
EP
2
H_1
59_R
EP
3
H_1
60_R
EP
1
H_1
60_R
EP
2
H_1
60_R
EP
3
H_1
61_R
EP
1
H_1
61_R
EP
2
H_1
61_R
EP
3
H_1
62_R
EP
1
H_1
62_R
EP
2
H_1
62_R
EP
3
Replicate
0
20
40
60
80
100
Peak
Are
a Pe
rcen
tage
y13 - 1426.6696+ y9 - 1024.5310+ y7 - 854.4254+ y6 - 691.3621+y4 - 477.2304+ y3 - 363.1874+
Libr
ary
D_1
02_R
EP
1
D_1
02_R
EP
2
D_1
02_R
EP
3
D_1
03_R
EP
1
D_1
03_R
EP
2
D_1
03_R
EP
3
D_1
08_R
EP
1
D_1
08_R
EP
2
D_1
08_R
EP
3
D_1
38_R
EP
1
D_1
38_R
EP
2
D_1
38_R
EP
3
D_1
54_R
EP
1
D_1
54_R
EP
2
D_1
54_R
EP
3
D_1
72_R
EP
1
D_1
72_R
EP
2
D_1
72_R
EP
3
D_1
96_R
EP
1
D_1
96_R
EP
2
D_1
96_R
EP
3
H_1
46_R
EP
1
H_1
46_R
EP
2
H_1
46_R
EP
3
H_1
47_R
EP
1
H_1
47_R
EP
2
H_1
47_R
EP
3
H_1
48_R
EP
1
H_1
48_R
EP
2
H_1
48_R
EP
3
H_1
59_R
EP
1
H_1
59_R
EP
2
H_1
59_R
EP
3
H_1
60_R
EP
1
H_1
60_R
EP
2
H_1
60_R
EP
3
H_1
61_R
EP
1
H_1
61_R
EP
2
H_1
61_R
EP
3
H_1
62_R
EP
1
H_1
62_R
EP
2
H_1
62_R
EP
3
Replicate
0
20
40
60
80
100
Peak
Are
a Pe
rcen
tage
y13 - 1426.6696+ y9 - 1024.5310+ y7 - 854.4254+ y6 - 691.3621+y4 - 477.2304+ y3 - 363.1874+
D_1
02_R
EP
1
D_1
02_R
EP
2
D_1
02_R
EP
3
D_1
03_R
EP
1
D_1
03_R
EP
2
D_1
03_R
EP
3
D_1
08_R
EP
1
D_1
08_R
EP
2
D_1
08_R
EP
3
D_1
38_R
EP
1
D_1
38_R
EP
2
D_1
38_R
EP
3
D_1
54_R
EP
1
D_1
54_R
EP
2
D_1
54_R
EP
3
D_1
72_R
EP
1
D_1
72_R
EP
2
D_1
72_R
EP
3
D_1
96_R
EP
1
D_1
96_R
EP
2
D_1
96_R
EP
3
H_1
46_R
EP
1
H_1
46_R
EP
2
H_1
46_R
EP
3
H_1
47_R
EP
1
H_1
47_R
EP
2
H_1
47_R
EP
3
H_1
48_R
EP
1
H_1
48_R
EP
2
H_1
48_R
EP
3
H_1
59_R
EP
1
H_1
59_R
EP
2
H_1
59_R
EP
3
H_1
60_R
EP
1
H_1
60_R
EP
2
H_1
60_R
EP
3
H_1
61_R
EP
1
H_1
61_R
EP
2
H_1
61_R
EP
3
H_1
62_R
EP
1
H_1
62_R
EP
2
H_1
62_R
EP
3
Replicate
16.5
17.0
17.5
18.0
Ret
entio
n Ti
me
y13 - 1426.6696+ y9 - 1024.5310+ y7 - 854.4254+ y6 - 691.3621+y4 - 477.2304+ y3 - 363.1874+
D_1
02_R
EP
1
D_1
02_R
EP
2
D_1
02_R
EP
3
D_1
03_R
EP
1
D_1
03_R
EP
2
D_1
03_R
EP
3
D_1
08_R
EP
1
D_1
08_R
EP
2
D_1
08_R
EP
3
D_1
38_R
EP
1
D_1
38_R
EP
2
D_1
38_R
EP
3
D_1
54_R
EP
1
D_1
54_R
EP
2
D_1
54_R
EP
3
D_1
72_R
EP
1
D_1
72_R
EP
2
D_1
72_R
EP
3
D_1
96_R
EP
1
D_1
96_R
EP
2
D_1
96_R
EP
3
H_1
46_R
EP
1
H_1
46_R
EP
2
H_1
46_R
EP
3
H_1
47_R
EP
1
H_1
47_R
EP
2
H_1
47_R
EP
3
H_1
48_R
EP
1
H_1
48_R
EP
2
H_1
48_R
EP
3
H_1
59_R
EP
1
H_1
59_R
EP
2
H_1
59_R
EP
3
H_1
60_R
EP
1
H_1
60_R
EP
2
H_1
60_R
EP
3
H_1
61_R
EP
1
H_1
61_R
EP
2
H_1
61_R
EP
3
H_1
62_R
EP
1
H_1
62_R
EP
2
H_1
62_R
EP
3
Replicate
16.5
17.0
17.5
18.0
Ret
entio
n Ti
me
y13 - 1426.6696+ y9 - 1024.5310+ y7 - 854.4254+ y6 - 691.3621+y4 - 477.2304+ y3 - 363.1874+
LQTEGDGIYTLNSEK
Haptoglobin Li
brar
y
D_1
02_R
EP
1
D_1
02_R
EP
2
D_1
02_R
EP
3
D_1
03_R
EP
1
D_1
03_R
EP
2
D_1
03_R
EP
3
D_1
08_R
EP
1
D_1
08_R
EP
2
D_1
08_R
EP
3
D_1
38_R
EP
1
D_1
38_R
EP
2
D_1
38_R
EP
3
D_1
54_R
EP
1
D_1
54_R
EP
2
D_1
54_R
EP
3
D_1
72_R
EP
1
D_1
72_R
EP
2
D_1
72_R
EP
3
D_1
96_R
EP
1
D_1
96_R
EP
2
D_1
96_R
EP
3
H_1
46_R
EP
1
H_1
46_R
EP
2
H_1
46_R
EP
3
H_1
47_R
EP
1
H_1
47_R
EP
2
H_1
47_R
EP
3
H_1
48_R
EP
1
H_1
48_R
EP
2
H_1
48_R
EP
3
H_1
59_R
EP
1
H_1
59_R
EP
2
H_1
59_R
EP
3
H_1
60_R
EP
1
H_1
60_R
EP
2
H_1
60_R
EP
3
H_1
61_R
EP
1
H_1
61_R
EP
2
H_1
61_R
EP
3
H_1
62_R
EP
1
H_1
62_R
EP
2
H_1
62_R
EP
3
Replicate
0
2
4
6
8
Peak
Are
a (1
0^6)
y13 - 1426.6696+ y9 - 1024.5310+ y7 - 854.4254+ y6 - 691.3621+ y4 - 477.2304+y3 - 363.1874+
Libr
ary
D_1
02_R
EP
1
D_1
02_R
EP
2
D_1
02_R
EP
3
D_1
03_R
EP
1
D_1
03_R
EP
2
D_1
03_R
EP
3
D_1
08_R
EP
1
D_1
08_R
EP
2
D_1
08_R
EP
3
D_1
38_R
EP
1
D_1
38_R
EP
2
D_1
38_R
EP
3
D_1
54_R
EP
1
D_1
54_R
EP
2
D_1
54_R
EP
3
D_1
72_R
EP
1
D_1
72_R
EP
2
D_1
72_R
EP
3
D_1
96_R
EP
1
D_1
96_R
EP
2
D_1
96_R
EP
3
H_1
46_R
EP
1
H_1
46_R
EP
2
H_1
46_R
EP
3
H_1
47_R
EP
1
H_1
47_R
EP
2
H_1
47_R
EP
3
H_1
48_R
EP
1
H_1
48_R
EP
2
H_1
48_R
EP
3
H_1
59_R
EP
1
H_1
59_R
EP
2
H_1
59_R
EP
3
H_1
60_R
EP
1
H_1
60_R
EP
2
H_1
60_R
EP
3
H_1
61_R
EP
1
H_1
61_R
EP
2
H_1
61_R
EP
3
H_1
62_R
EP
1
H_1
62_R
EP
2
H_1
62_R
EP
3
Replicate
0
2
4
6
8
Peak
Are
a (1
0^6)
y13 - 1426.6696+ y9 - 1024.5310+ y7 - 854.4254+ y6 - 691.3621+ y4 - 477.2304+y3 - 363.1874+
LQTEGDGIYTLNSEK Li
brar
y
D_1
02_R
EP
1
D_1
02_R
EP
2
D_1
02_R
EP
3
D_1
03_R
EP
1
D_1
03_R
EP
2
D_1
03_R
EP
3
D_1
08_R
EP
1
D_1
08_R
EP
2
D_1
08_R
EP
3
D_1
38_R
EP
1
D_1
38_R
EP
2
D_1
38_R
EP
3
D_1
54_R
EP
1
D_1
54_R
EP
2
D_1
54_R
EP
3
D_1
72_R
EP
1
D_1
72_R
EP
2
D_1
72_R
EP
3
D_1
96_R
EP
1
D_1
96_R
EP
2
D_1
96_R
EP
3
H_1
46_R
EP
1
H_1
46_R
EP
2
H_1
46_R
EP
3
H_1
47_R
EP
1
H_1
47_R
EP
2
H_1
47_R
EP
3
H_1
48_R
EP
1
H_1
48_R
EP
2
H_1
48_R
EP
3
H_1
59_R
EP
1
H_1
59_R
EP
2
H_1
59_R
EP
3
H_1
60_R
EP
1
H_1
60_R
EP
2
H_1
60_R
EP
3
H_1
61_R
EP
1
H_1
61_R
EP
2
H_1
61_R
EP
3
H_1
62_R
EP
1
H_1
62_R
EP
2
H_1
62_R
EP
3
Replicate
0.0
0.5
1.0
1.5
2.0
Peak
Are
a (1
0^6)
y7 - 757.4818+ y6 - 658.4134+ y5 - 543.3865+ y4 - 430.3024+
Libr
ary
D_1
02_R
EP
1
D_1
02_R
EP
2
D_1
02_R
EP
3
D_1
03_R
EP
1
D_1
03_R
EP
2
D_1
03_R
EP
3
D_1
08_R
EP
1
D_1
08_R
EP
2
D_1
08_R
EP
3
D_1
38_R
EP
1
D_1
38_R
EP
2
D_1
38_R
EP
3
D_1
54_R
EP
1
D_1
54_R
EP
2
D_1
54_R
EP
3
D_1
72_R
EP
1
D_1
72_R
EP
2
D_1
72_R
EP
3
D_1
96_R
EP
1
D_1
96_R
EP
2
D_1
96_R
EP
3
H_1
46_R
EP
1
H_1
46_R
EP
2
H_1
46_R
EP
3
H_1
47_R
EP
1
H_1
47_R
EP
2
H_1
47_R
EP
3
H_1
48_R
EP
1
H_1
48_R
EP
2
H_1
48_R
EP
3
H_1
59_R
EP
1
H_1
59_R
EP
2
H_1
59_R
EP
3
H_1
60_R
EP
1
H_1
60_R
EP
2
H_1
60_R
EP
3
H_1
61_R
EP
1
H_1
61_R
EP
2
H_1
61_R
EP
3
H_1
62_R
EP
1
H_1
62_R
EP
2
H_1
62_R
EP
3
Replicate
0.0
0.5
1.0
1.5
2.0
Peak
Are
a (1
0^6)
y7 - 757.4818+ y6 - 658.4134+ y5 - 543.3865+ y4 - 430.3024+
SVVDIGLIK
Mitochondrial 39S ribosomal protein L9D
_102
_RE
P1
D_1
02_R
EP
2
D_1
02_R
EP
3
D_1
03_R
EP
1
D_1
03_R
EP
2
D_1
03_R
EP
3
D_1
08_R
EP
1
D_1
08_R
EP
2
D_1
08_R
EP
3
D_1
38_R
EP
1
D_1
38_R
EP
2
D_1
38_R
EP
3
D_1
54_R
EP
1
D_1
54_R
EP
2
D_1
54_R
EP
3
D_1
72_R
EP
1
D_1
72_R
EP
2
D_1
72_R
EP
3
D_1
96_R
EP
1
D_1
96_R
EP
2
D_1
96_R
EP
3
H_1
46_R
EP
1
H_1
46_R
EP
2
H_1
46_R
EP
3
H_1
47_R
EP
1
H_1
47_R
EP
2
H_1
47_R
EP
3
H_1
48_R
EP
1
H_1
48_R
EP
2
H_1
48_R
EP
3
H_1
59_R
EP
1
H_1
59_R
EP
2
H_1
59_R
EP
3
H_1
60_R
EP
1
H_1
60_R
EP
2
H_1
60_R
EP
3
H_1
61_R
EP
1
H_1
61_R
EP
2
H_1
61_R
EP
3
H_1
62_R
EP
1
H_1
62_R
EP
2
H_1
62_R
EP
3
Replicate
0
20
40
60
80
100
Peak
Are
a Pe
rcen
tage
y8 - 930.4944+ y7 - 744.4151+ y6 - 673.3780+ y5 - 616.3566+ y4 - 545.3194+y3 - 474.2823+
D_1
02_R
EP
1
D_1
02_R
EP
2
D_1
02_R
EP
3
D_1
03_R
EP
1
D_1
03_R
EP
2
D_1
03_R
EP
3
D_1
08_R
EP
1
D_1
08_R
EP
2
D_1
08_R
EP
3
D_1
38_R
EP
1
D_1
38_R
EP
2
D_1
38_R
EP
3
D_1
54_R
EP
1
D_1
54_R
EP
2
D_1
54_R
EP
3
D_1
72_R
EP
1
D_1
72_R
EP
2
D_1
72_R
EP
3
D_1
96_R
EP
1
D_1
96_R
EP
2
D_1
96_R
EP
3
H_1
46_R
EP
1
H_1
46_R
EP
2
H_1
46_R
EP
3
H_1
47_R
EP
1
H_1
47_R
EP
2
H_1
47_R
EP
3
H_1
48_R
EP
1
H_1
48_R
EP
2
H_1
48_R
EP
3
H_1
59_R
EP
1
H_1
59_R
EP
2
H_1
59_R
EP
3
H_1
60_R
EP
1
H_1
60_R
EP
2
H_1
60_R
EP
3
H_1
61_R
EP
1
H_1
61_R
EP
2
H_1
61_R
EP
3
H_1
62_R
EP
1
H_1
62_R
EP
2
H_1
62_R
EP
3
Replicate
0
20
40
60
80
100
Peak
Are
a Pe
rcen
tage
y8 - 930.4944+ y7 - 744.4151+ y6 - 673.3780+ y5 - 616.3566+ y4 - 545.3194+y3 - 474.2823+
D_1
02_R
EP
1
D_1
02_R
EP
2
D_1
02_R
EP
3
D_1
03_R
EP
1
D_1
03_R
EP
2
D_1
03_R
EP
3
D_1
08_R
EP
1
D_1
08_R
EP
2
D_1
08_R
EP
3
D_1
38_R
EP
1
D_1
38_R
EP
2
D_1
38_R
EP
3
D_1
54_R
EP
1
D_1
54_R
EP
2
D_1
54_R
EP
3
D_1
72_R
EP
1
D_1
72_R
EP
2
D_1
72_R
EP
3
D_1
96_R
EP
1
D_1
96_R
EP
2
D_1
96_R
EP
3
H_1
46_R
EP
1
H_1
46_R
EP
2
H_1
46_R
EP
3
H_1
47_R
EP
1
H_1
47_R
EP
2
H_1
47_R
EP
3
H_1
48_R
EP
1
H_1
48_R
EP
2
H_1
48_R
EP
3
H_1
59_R
EP
1
H_1
59_R
EP
2
H_1
59_R
EP
3
H_1
60_R
EP
1
H_1
60_R
EP
2
H_1
60_R
EP
3
H_1
61_R
EP
1
H_1
61_R
EP
2
H_1
61_R
EP
3
H_1
62_R
EP
1
H_1
62_R
EP
2
H_1
62_R
EP
3
Replicate
19.520.020.521.021.522.022.5
Ret
entio
n Ti
me
y8 - 930.4944+ y7 - 744.4151+ y6 - 673.3780+ y5 - 616.3566+ y4 - 545.3194+y3 - 474.2823+
D_1
02_R
EP
1
D_1
02_R
EP
2
D_1
02_R
EP
3
D_1
03_R
EP
1
D_1
03_R
EP
2
D_1
03_R
EP
3
D_1
08_R
EP
1
D_1
08_R
EP
2
D_1
08_R
EP
3
D_1
38_R
EP
1
D_1
38_R
EP
2
D_1
38_R
EP
3
D_1
54_R
EP
1
D_1
54_R
EP
2
D_1
54_R
EP
3
D_1
72_R
EP
1
D_1
72_R
EP
2
D_1
72_R
EP
3
D_1
96_R
EP
1
D_1
96_R
EP
2
D_1
96_R
EP
3
H_1
46_R
EP
1
H_1
46_R
EP
2
H_1
46_R
EP
3
H_1
47_R
EP
1
H_1
47_R
EP
2
H_1
47_R
EP
3
H_1
48_R
EP
1
H_1
48_R
EP
2
H_1
48_R
EP
3
H_1
59_R
EP
1
H_1
59_R
EP
2
H_1
59_R
EP
3
H_1
60_R
EP
1
H_1
60_R
EP
2
H_1
60_R
EP
3
H_1
61_R
EP
1
H_1
61_R
EP
2
H_1
61_R
EP
3
H_1
62_R
EP
1
H_1
62_R
EP
2
H_1
62_R
EP
3
Replicate
19.520.020.521.021.522.022.5
Ret
entio
n Ti
me
y8 - 930.4944+ y7 - 744.4151+ y6 - 673.3780+ y5 - 616.3566+ y4 - 545.3194+y3 - 474.2823+
CSSLLWAGAAWLR
Mitochondrial 39S ribosomal protein L9D
_102
_RE
P1
D_1
02_R
EP
2
D_1
02_R
EP
3
D_1
03_R
EP
1
D_1
03_R
EP
2
D_1
03_R
EP
3
D_1
08_R
EP
1
D_1
08_R
EP
2
D_1
08_R
EP
3
D_1
38_R
EP
1
D_1
38_R
EP
2
D_1
38_R
EP
3
D_1
54_R
EP
1
D_1
54_R
EP
2
D_1
54_R
EP
3
D_1
72_R
EP
1
D_1
72_R
EP
2
D_1
72_R
EP
3
D_1
96_R
EP
1
D_1
96_R
EP
2
D_1
96_R
EP
3
H_1
46_R
EP
1
H_1
46_R
EP
2
H_1
46_R
EP
3
H_1
47_R
EP
1
H_1
47_R
EP
2
H_1
47_R
EP
3
H_1
48_R
EP
1
H_1
48_R
EP
2
H_1
48_R
EP
3
H_1
59_R
EP
1
H_1
59_R
EP
2
H_1
59_R
EP
3
H_1
60_R
EP
1
H_1
60_R
EP
2
H_1
60_R
EP
3
H_1
61_R
EP
1
H_1
61_R
EP
2
H_1
61_R
EP
3
H_1
62_R
EP
1
H_1
62_R
EP
2
H_1
62_R
EP
3
Replicate
020406080
100120140
Peak
Are
a (1
0^3)
y8 - 930.4944+ y7 - 744.4151+ y6 - 673.3780+ y5 - 616.3566+ y4 - 545.3194+y3 - 474.2823+
D_1
02_R
EP
1
D_1
02_R
EP
2
D_1
02_R
EP
3
D_1
03_R
EP
1
D_1
03_R
EP
2
D_1
03_R
EP
3
D_1
08_R
EP
1
D_1
08_R
EP
2
D_1
08_R
EP
3
D_1
38_R
EP
1
D_1
38_R
EP
2
D_1
38_R
EP
3
D_1
54_R
EP
1
D_1
54_R
EP
2
D_1
54_R
EP
3
D_1
72_R
EP
1
D_1
72_R
EP
2
D_1
72_R
EP
3
D_1
96_R
EP
1
D_1
96_R
EP
2
D_1
96_R
EP
3
H_1
46_R
EP
1
H_1
46_R
EP
2
H_1
46_R
EP
3
H_1
47_R
EP
1
H_1
47_R
EP
2
H_1
47_R
EP
3
H_1
48_R
EP
1
H_1
48_R
EP
2
H_1
48_R
EP
3
H_1
59_R
EP
1
H_1
59_R
EP
2
H_1
59_R
EP
3
H_1
60_R
EP
1
H_1
60_R
EP
2
H_1
60_R
EP
3
H_1
61_R
EP
1
H_1
61_R
EP
2
H_1
61_R
EP
3
H_1
62_R
EP
1
H_1
62_R
EP
2
H_1
62_R
EP
3
Replicate
020406080
100120140
Peak
Are
a (1
0^3)
y8 - 930.4944+ y7 - 744.4151+ y6 - 673.3780+ y5 - 616.3566+ y4 - 545.3194+y3 - 474.2823+
D_1
02_R
EP
1
D_1
02_R
EP
2
D_1
02_R
EP
3
D_1
03_R
EP
1
D_1
03_R
EP
2
D_1
03_R
EP
3
D_1
08_R
EP
1
D_1
08_R
EP
2
D_1
08_R
EP
3
D_1
38_R
EP
1
D_1
38_R
EP
2
D_1
38_R
EP
3
D_1
54_R
EP
1
D_1
54_R
EP
2
D_1
54_R
EP
3
D_1
72_R
EP
1
D_1
72_R
EP
2
D_1
72_R
EP
3
D_1
96_R
EP
1
D_1
96_R
EP
2
D_1
96_R
EP
3
H_1
46_R
EP
1
H_1
46_R
EP
2
H_1
46_R
EP
3
H_1
47_R
EP
1
H_1
47_R
EP
2
H_1
47_R
EP
3
H_1
48_R
EP
1
H_1
48_R
EP
2
H_1
48_R
EP
3
H_1
59_R
EP
1
H_1
59_R
EP
2
H_1
59_R
EP
3
H_1
60_R
EP
1
H_1
60_R
EP
2
H_1
60_R
EP
3
H_1
61_R
EP
1
H_1
61_R
EP
2
H_1
61_R
EP
3
H_1
62_R
EP
1
H_1
62_R
EP
2
H_1
62_R
EP
3
Replicate
0
200
400
600
800
Peak
Are
a (1
0^3)
y9 - 961.5942+ y8 - 862.5257+ y6 - 664.3889+ y5 - 593.3518+ y4 - 496.2990+y3 - 359.2401+
D_1
02_R
EP
1
D_1
02_R
EP
2
D_1
02_R
EP
3
D_1
03_R
EP
1
D_1
03_R
EP
2
D_1
03_R
EP
3
D_1
08_R
EP
1
D_1
08_R
EP
2
D_1
08_R
EP
3
D_1
38_R
EP
1
D_1
38_R
EP
2
D_1
38_R
EP
3
D_1
54_R
EP
1
D_1
54_R
EP
2
D_1
54_R
EP
3
D_1
72_R
EP
1
D_1
72_R
EP
2
D_1
72_R
EP
3
D_1
96_R
EP
1
D_1
96_R
EP
2
D_1
96_R
EP
3
H_1
46_R
EP
1
H_1
46_R
EP
2
H_1
46_R
EP
3
H_1
47_R
EP
1
H_1
47_R
EP
2
H_1
47_R
EP
3
H_1
48_R
EP
1
H_1
48_R
EP
2
H_1
48_R
EP
3
H_1
59_R
EP
1
H_1
59_R
EP
2
H_1
59_R
EP
3
H_1
60_R
EP
1
H_1
60_R
EP
2
H_1
60_R
EP
3
H_1
61_R
EP
1
H_1
61_R
EP
2
H_1
61_R
EP
3
H_1
62_R
EP
1
H_1
62_R
EP
2
H_1
62_R
EP
3
Replicate
0
200
400
600
800
Peak
Are
a (1
0^3)
y9 - 961.5942+ y8 - 862.5257+ y6 - 664.3889+ y5 - 593.3518+ y4 - 496.2990+y3 - 359.2401+
SVVDIGLIK
CSSLLWAGAAWLR
CSSLLWAGAAWLRH_159_REP1
20.0 20.5 21.0 21.5
Retention Time
0
1000
2000
3000
4000
5000
Inte
nsity
20.7
21.6
20.4 20.6 20.8 21.0 21.2 21.4
Retention Time
0
20
40
60
80
100
120
140
Inte
nsity
21.021.4
D_138_REP3
18 19 20 21 22 23 24 25
Retention Time
0100200300400500600700800900
Inte
nsity
21.0
22.5
23.119.4
22.8
20.0
19.6
D_1
02_R
EP
1
D_1
03_R
EP
1
D_1
08_R
EP
1
D_1
38_R
EP
1
D_1
54_R
EP
1
D_1
72_R
EP
1
D_1
96_R
EP
1
H_1
46_R
EP
1
H_1
47_R
EP
1
H_1
48_R
EP
1
H_1
59_R
EP
1
H_1
60_R
EP
1
H_1
61_R
EP
1
H_1
62_R
EP
1
D_1
02_R
EP
2
D_1
03_R
EP
2
D_1
08_R
EP
2
D_1
38_R
EP
2
D_1
54_R
EP
2
D_1
72_R
EP
2
D_1
96_R
EP
2
H_1
46_R
EP
2
H_1
47_R
EP
2
H_1
48_R
EP
2
H_1
59_R
EP
2
H_1
60_R
EP
2
H_1
61_R
EP
2
H_1
62_R
EP
2
D_1
02_R
EP
3
D_1
03_R
EP
3
D_1
08_R
EP
3
D_1
38_R
EP
3
D_1
54_R
EP
3
D_1
72_R
EP
3
D_1
96_R
EP
3
H_1
46_R
EP
3
H_1
47_R
EP
3
H_1
48_R
EP
3
H_1
59_R
EP
3
H_1
60_R
EP
3
H_1
61_R
EP
3
H_1
62_R
EP
3
Replicate
19.520.020.521.021.522.022.5
Ret
entio
n Ti
me
y8 - 930.4944+ y7 - 744.4151+ y6 - 673.3780+ y5 - 616.3566+ y4 - 545.3194+y3 - 474.2823+
D_1
02_R
EP
1
D_1
03_R
EP
1
D_1
08_R
EP
1
D_1
38_R
EP
1
D_1
54_R
EP
1
D_1
72_R
EP
1
D_1
96_R
EP
1
H_1
46_R
EP
1
H_1
47_R
EP
1
H_1
48_R
EP
1
H_1
59_R
EP
1
H_1
60_R
EP
1
H_1
61_R
EP
1
H_1
62_R
EP
1
D_1
02_R
EP
2
D_1
03_R
EP
2
D_1
08_R
EP
2
D_1
38_R
EP
2
D_1
54_R
EP
2
D_1
72_R
EP
2
D_1
96_R
EP
2
H_1
46_R
EP
2
H_1
47_R
EP
2
H_1
48_R
EP
2
H_1
59_R
EP
2
H_1
60_R
EP
2
H_1
61_R
EP
2
H_1
62_R
EP
2
D_1
02_R
EP
3
D_1
03_R
EP
3
D_1
08_R
EP
3
D_1
38_R
EP
3
D_1
54_R
EP
3
D_1
72_R
EP
3
D_1
96_R
EP
3
H_1
46_R
EP
3
H_1
47_R
EP
3
H_1
48_R
EP
3
H_1
59_R
EP
3
H_1
60_R
EP
3
H_1
61_R
EP
3
H_1
62_R
EP
3
Replicate
19.520.020.521.021.522.022.5
Ret
entio
n Ti
me
y8 - 930.4944+ y7 - 744.4151+ y6 - 673.3780+ y5 - 616.3566+ y4 - 545.3194+y3 - 474.2823+
Truncated and Missing PeaksTGTNLMDFLSR
Deciphering the UnexpectedA
1A
3A
4A
5A
6A
7A
8A
9A
10 A11
A12 B
1B
2B
3B
4B
5B
6B
7B
8B
10 B11
B12 C
1C
2C
3C
4C
5C
6C
7C
8C
9C
10C
11C
12 D1
D2
D4
D5
D6
D7
D8
D9
D10
D11
D12 E
1E
2E
3E
4E
5E
6E
7E
8E
9E
10 E11
E12 F1 F2 F3 F4 F5 F6 F7 F8 F9 F1
0F1
1F1
2G
1G
2G
3G
4G
5G
6G
7G
8G
9G
10G
11G
12
Replicate
19.2
19.4
19.6
19.8
20.0
20.2
20.4
20.6
20.8
21.0
Ret
entio
n Ti
me
YGQTIRPICLPCTEGTTR - 708.3506+++ YGQTIRPICLPCTEGTTR - 711.6867+++ (heavy)
A1
A3
A4
A5
A6
A7
A8
A9
A10 A11
A12 B
1B
2B
3B
4B
5B
6B
7B
8B
10 B11
B12 C
1C
2C
3C
4C
5C
6C
7C
8C
9C
10C
11C
12 D1
D2
D4
D5
D6
D7
D8
D9
D10
D11
D12 E
1E
2E
3E
4E
5E
6E
7E
8E
9E
10 E11
E12 F1 F2 F3 F4 F5 F6 F7 F8 F9 F1
0F1
1F1
2G
1G
2G
3G
4G
5G
6G
7G
8G
9G
10G
11G
12
Replicate
19.2
19.4
19.6
19.8
20.0
20.2
20.4
20.6
20.8
21.0
Ret
entio
n Ti
me
YGQTIRPICLPCTEGTTR - 708.3506+++ YGQTIRPICLPCTEGTTR - 711.6867+++ (heavy) YGQTIRPICLPCTEGTTR
17 to 22-Feb-2012 16 to 17-Mar-2012
Aligned by iRTA
1A
3A
4A
5A
6A
7A
8A
9A
10 A11
A12 B
1B
2B
3B
4B
5B
6B
7B
8B
10 B11
B12 C
1C
2C
3C
4C
5C
6C
7C
8C
9C
10C
11C
12 D1
D2
D4
D5
D6
D7
D8
D9
D10
D11
D12 E
1E
2E
3E
4E
5E
6E
7E
8E
9E
10 E11
E12 F1 F2 F3 F4 F5 F6 F7 F8 F9 F1
0F1
1F1
2G
1G
2G
3G
4G
5G
6G
7G
8G
9G
10G
11G
12
Replicate
37
38
39
40
41
42
43
iRT
(Hut
tenh
ain)
Sco
re
YGQTIRPICLPCTEGTTR - 708.3506+++ YGQTIRPICLPCTEGTTR - 711.6867+++ (heavy)
A1
A3
A4
A5
A6
A7
A8
A9
A10 A11
A12 B
1B
2B
3B
4B
5B
6B
7B
8B
10 B11
B12 C
1C
2C
3C
4C
5C
6C
7C
8C
9C
10C
11C
12 D1
D2
D4
D5
D6
D7
D8
D9
D10
D11
D12 E
1E
2E
3E
4E
5E
6E
7E
8E
9E
10 E11
E12 F1 F2 F3 F4 F5 F6 F7 F8 F9 F1
0F1
1F1
2G
1G
2G
3G
4G
5G
6G
7G
8G
9G
10G
11G
12
Replicate
37
38
39
40
41
42
43
iRT
(Hut
tenh
ain)
Sco
re
YGQTIRPICLPCTEGTTR - 708.3506+++ YGQTIRPICLPCTEGTTR - 711.6867+++ (heavy) YGQTIRPICLPCTEGTTR
Integrating with External Tools GeneSpring & Skyline Automation Tool
(Agilent) MSstats (Purdue) – Grouped study analysis QuaSAR (Broad Institute) – Response curves,
LOD, LOQ MS1 Probe (Buck Institute) – MS1 quant.
statistics TP28 – Alexandria D’Souza
SRM Collider (IMSB) – Interference probability calculator
External Tool MSstats
Replicate Annotations
Custom Reports
Analysis of reports with R – MSstatsDownstream Analysis with Statistical Tools
Skyline Automation with SkylineRunner
MS/MS Spectral Library Sources Global Proteome Machine NIST Peptide Atlas
Build your own from peptide search results Mascot MaxQuant Andromeda Morpheus Myrimatch / IDPicker OMSSA PRIDE XML Protein Pilot Protein Prospector
Proteome Discoverer (MSF) Scaffold – mzIdentML / MGF Spectrum Mill TPP – pepXML / mzXML files –
Peptide Atlas Waters Mse X! Tandem
Integrating Great Ideas (iRT)
Integrating Great Ideas (mProphet)
TP 499 - MacLean
Skyline Team Nick Shulman
Don Marsh
Kaipo Tamura
Danny Broudy Trevor Killeen
Vagish Sharma
Josh Eckels
Greg Taylor
Jarrett Egertson Dario Amodei
Collaborators: U. of Wa.
Eva Baker Jarrett Egertson Jimmy Eng Andrew Stergachis
Biognosys Lukas Reiter Oliver Rinner Claudia Escher
Broad Institute Sue Abbatiello Steve Carr Jake Jaffe D. R. Mani
Buck Institute Birgit Schilling Matthew Rardin Brad Gibson
Duke Will Thompson Arthur Moseley
IMSB Rudolph Aebersold Christina Ludwig Olga Schubert Hannes Röst Lucia Espona Pernas
Purdue Veavi Chang Meena Choi Olga Vitek
Stanford Dario Amodei Parag Mallick
Vanderbilt Matthew Chambers Daniel Liebler David Tabb
Instrument Vendor Support Agilent
Technologies Christine Miller Juli Salcedo Shripad Torvi Yinghang Yang
Bruker Carsten Baessmann Marius Kallhardt Stephanie Kaspar
AB Sciex David Cox Christie Hunter Brent Lefebvre
Thermo-Scientific Markus Kellmann Andreas Kuehn Vlad Zabrouskov
Waters Laurence Firth James Langridge Roy Martin Kieran Neeson Keith Richards