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What is the Spectrum Mill MS Proteomics Workbench?
Agilent G2721AA Spectrum Mill
MS Proteomics Workbench
Quick Start Guide
A guide to the Spectrum Mill workbench
Use this reference for your first steps with the Spectrum Mill workbench.
The Spectrum Mill software is a collection of tools for high-
throughput processing of MS and MS/MS spectra to provide protein
and peptide identifications and relative quantitation. The
Spectrum Mill workbench can do in minutes what could take you
hours or days to do manually.
This software includes bioinformatics tools for:
• Spectral preprocessing for MS/MS and MS-only data, including
spectral quality filtering and precursor charge state assignment
for MS/MS spectra
• Protein database search, including searches for both MS/MS and
MS-only spectra
• Data review, validation, and comparison for large data sets,
including those from two-dimensional LC/MS/MS analyses and
differential expression studies
• De novo sequencing
• Identification of mutations, post-translational modifications,
and chemical modifications
• Differential expression quantitation
The Spectrum Mill workbench accepts Agilent data, as well as data
from other vendors' mass spectrometers. The software executes on
a web server, and you access the program via a web browser
window.
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Table of Contents
Related Manuals for Agilent Technologies Spectrum Mill MS G2721AA
Summary of Contents for Agilent Technologies Spectrum Mill MS G2721AA
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Page 1: Quick Start Guide
• Differential expression quantitation The Spectrum Mill workbench accepts Agilent data, as well as data from other vendors’ mass spectrometers. The software executes on a web server, and you access the program via a web browser window. Agilent Technologies... - Page 2 New features in Spectrum Mill workbench version A.03.03 The following are key changes to the user interface and functionality: Support for more Now supported: instruments and • Agilent Q-TOF data types • Electron transfer dissociation (ETD) data from Agilent ion trap •...
- Page 3 Protein/Peptide Spectrum Viewer support for c- and z-ions, which are prominent in ETD spectra Summary Retention time in peptide summaries Fragmentation mode in peptide summaries, so you can ascertain the mode for each spectrum (for example, CID versus ETD) Enhanced protein grouping based upon shared peptides Ability to align sequences of proteins within a protein group Ability to sort and filter by peptide pI New capability in the Spectrum Viewer to type your own sequence and have it add...
- Page 4 The ability to store iTRAQ correction factors for multiple iTRAQ batches and to apply them to the iTRAQ calculations for specific data sets Expanded capabilities for Peptide Selector Updated instrument selection lists The default Spectrum Mill font size is now “smaller”, so you may use the default “medium”...
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Page 5: Spectrum Mill Manuals And Online Help
Spectrum Mill manuals and online help Scientists Familiarization Guide Follow step-by-step instructions to process example data. Application Guide Learn details to use the software. Online Help Consult the online help for in-depth information not given in the Application Guide. To access help, click links on the home page, Help buttons on Spectrum Mill forms, or blue dividing bars on Spectrum Mill forms. -
Page 6: Setting Up The Spectrum Mill Server
Setting up the Spectrum Mill server See the Spectrum Mill MS Proteomics Workbench Installation Guide. If you wish to update from a previous version of the Spectrum Mill workbench, see Chapter 2. Note that the server name cannot have an underscore. Setting up your client PC Operating system Check that you have one of the following:... - Page 7 Stored pages Disable caching. 1 With IE 6.0 or IE 7.0, select Tools > Internet Options..2 Click the General tab. 3 Do one of the following: • If you have IE 6.0, under Temporary internet files, click the Settings... button.
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Page 8: Running The Spectrum Mill Workbench From The Client
Running the Spectrum Mill workbench from the client Open Internet Explorer and point to the URL for the Spectrum Mill server. This has the general format server/millhome.htm. If you are not sure of the URL, ask your system administrator. Setting up Agilent TOF analyses for use with the Spectrum Mill workbench When you set up data acquisition parameters on the Agilent TOF, it is critical that you establish analysis settings that are compatible with Spectrum Mill processing. -
Page 9: Spectrum Mill Modules
Spectrum Mill modules Tool type Module Function Application Guide Chapter(s) Spectral Data Extractor Extract spectra from data files. Processing depends 1, 6 preprocessing on data type and may include averaging, centroiding, filtering by quality, assigning precursor charge, and calculating spectral features. Interpretation MS/MS Search Search MS/MS spectra against databases. - Page 10 Tool type Module Function Application Guide Chapter(s) Utilities Tool Belt Use tools to speed processing, examine parameters and search statistics, stop processes, etc. Protein Databases Create indices for databases so searches run faster, create subset databases, etc. Build TIC Visualize how much of your sample has been interpreted.
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Page 11: Roadmap For Ms/Ms Data Processing
Roadmap for MS/MS data processing See Chapter 1 in the Application Guide for details. Start here. Preprocess spectral files (Data Set search parameters and run Collect and transfer data to Extractor page ) searches (MS/MS Search page ) Spectrum Mill server Summarize valid results Validate medium-quality results Validate highest-quality results... - Page 12 The following iterative search strategy for the Spectrum Mill workbench assumes that the goal is to identify as many proteins as possible. If your study does not require you to identify so many proteins, you may omit some steps (those between step 5 step 13).
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Page 13: Roadmap For Ms-Only Data Processing
Roadmap for MS-only data processing See Chapter 6 in the Application Guide for details. Start here. Preprocess Prepare and transfer spectral files Do you have QSTAR, or mass list files to (Data Extractor Agilent TOF or Trap data? Spectrum Mill page) server Set search parameters and run... -
Page 14: Modes For Protein/Peptide Summary
Modes for Protein/Peptide Summary See Chapter 2 in the Application Guide for details. If you want to: And you want results Then use this mode: Example application organized by: Validate results Peptides Peptide Manual review and validation of MS/MS search results, organized by peptide Proteins, then peptides Protein Summary... -
Page 15: Guide To Common Buttons
Guide to common buttons Any green button Initiates a process. Also saves form settings (except for data directory) until you close your web browser window. Save Settings Saves form settings (except for data directory) for future web browser sessions. Select Selects a data directory for this and subsequent forms. -
Page 16: Interpreting Scores From Ms/Ms Search
Interpreting scores from MS/MS Search The following definitions and guidelines will help you evaluate your MS/MS search results: Peptide score Score of an individual peptide. This score reflects the information content (amount of useful fragmentation) in the MS/MS spectrum. Protein score Score of the overall protein, calculated by adding scores of all the peptides detected for the protein •... - Page 17 Spectrum Mill Workbench Quick Start Guide...
- Page 18 Spectrum Mill Workbench Quick Start Guide...
- Page 19 Spectrum Mill Workbench Quick Start Guide...
- Page 20 In this Book The Quick Start Guide presents first steps to use the Spectrum Mill MS Proteomics Workbench. © Agilent Technologies, Inc. 2007 Printed in USA Fifth edition, June 2007 *G2721-90028* G2721-90028 Agilent Technologies...