2. Evotips:
(a) Disposable C-18 trap column, Evotips (Evosep Biosystems, Denmark).
(b) EV-1068 Rack holder (Evosep Biosystems, Denmark).
(c) 96 well Microtiter plate (96 well Conical Btm Pit Natural,
Thermo Fisher Scientific).
3. LC–MS instrumentation and software:
(a) Evosep One LC instrument, Evosep One system (Evosep
Biosystems, Denmark), and Evosep control software Axel
Semrau Chronos (Axel Semrau GmbH, Germany).
(b) C-18 nanoConnect LC column (8 cm column, ID
100 μm packed with 3 μm Reprocil, PepSep, Denmark)
with a Nano Source Emitter Stainless Steel Nano-bore
1/32 (Thermo Fisher Scientific, MA, USA).
(c) TripleTOF 6600 instrument (Sciex) equipped with a
NanoSpray III ion source (Sciex) and Mass spectrometer
acquisition software Analyst TF 1.7.1 (Sciex) and PeakView (Sciex).
2.7 Data Analysis
Workflow
1. Data sets for SEC-SWATH example:
(a) The mass spectrometry proteomic data have been deposited to the ProteomeXchange Consortium via the PRIDE
[25] partner repository with the data set identifier
PXD018033. Download the SWATH files: the
non-centroided *mzXML files from ProteomeXchange.
(b) Spectral assay library file and iRT file: Download the
adapted Pan Human library and the combined_iRT_CiRT_201804.TraML, which contains assays for the
retention time calibration (iRT) peptides [26] and
14 additional peptides of common lab-contaminants
from ProteomeXchange (data set identifier PXD018033).
2. Raw files were converted with ProteoWizard software (version
3.0.8851) http://proteowizard.sourceforge.net/projects.html
3. OpenSWATH library generation (Trans Proteomic Pipeline
5.0.0) https://sourceforge.net/projects/sashimi/files/latest/
download?source¼files
4. OpenSWATH workflow (OpenMS 2.4, 05.09.2019) https://
www.openms.de/download/openms-binaries/
5. PyProphet
(version
2.1.4.dev2)
https://github.com/
PyProphet/
6. TRIC: an automated alignment strategy for reproducible protein quantification in targeted proteomics. The TRIC package
is available under the following git repository: https://github.
System-Wide Profiling of Protein Complexes Via Size Exclusion. . .
275
(a) Disposable C-18 trap column, Evotips (Evosep Biosystems, Denmark).
(b) EV-1068 Rack holder (Evosep Biosystems, Denmark).
(c) 96 well Microtiter plate (96 well Conical Btm Pit Natural,
Thermo Fisher Scientific).
3. LC–MS instrumentation and software:
(a) Evosep One LC instrument, Evosep One system (Evosep
Biosystems, Denmark), and Evosep control software Axel
Semrau Chronos (Axel Semrau GmbH, Germany).
(b) C-18 nanoConnect LC column (8 cm column, ID
100 μm packed with 3 μm Reprocil, PepSep, Denmark)
with a Nano Source Emitter Stainless Steel Nano-bore
1/32 (Thermo Fisher Scientific, MA, USA).
(c) TripleTOF 6600 instrument (Sciex) equipped with a
NanoSpray III ion source (Sciex) and Mass spectrometer
acquisition software Analyst TF 1.7.1 (Sciex) and PeakView (Sciex).
2.7 Data Analysis
Workflow
1. Data sets for SEC-SWATH example:
(a) The mass spectrometry proteomic data have been deposited to the ProteomeXchange Consortium via the PRIDE
[25] partner repository with the data set identifier
PXD018033. Download the SWATH files: the
non-centroided *mzXML files from ProteomeXchange.
(b) Spectral assay library file and iRT file: Download the
adapted Pan Human library and the combined_iRT_CiRT_201804.TraML, which contains assays for the
retention time calibration (iRT) peptides [26] and
14 additional peptides of common lab-contaminants
from ProteomeXchange (data set identifier PXD018033).
2. Raw files were converted with ProteoWizard software (version
3.0.8851) http://proteowizard.sourceforge.net/projects.html
3. OpenSWATH library generation (Trans Proteomic Pipeline
5.0.0) https://sourceforge.net/projects/sashimi/files/latest/
download?source¼files
4. OpenSWATH workflow (OpenMS 2.4, 05.09.2019) https://
www.openms.de/download/openms-binaries/
5. PyProphet
(version
2.1.4.dev2)
https://github.com/
PyProphet/
6. TRIC: an automated alignment strategy for reproducible protein quantification in targeted proteomics. The TRIC package
is available under the following git repository: https://github.
System-Wide Profiling of Protein Complexes Via Size Exclusion. . .
275