The following article outlines the basic Scaffold workflow; which can be broken down into three main parts, see the diagram below:
Step 1. Generation of Raw Data: Samples are processed via LC- MS/MS. Raw spectra output files are generated most often in the proprietary format of the instrument manufacturer (Thermo result files are written with the
RAW extension for example).
Step 2. Peptide identification: Raw data files are loaded into a search engine application. Although X! Tandem comes bundled, Scaffold is not a search engine. There are various open source and commercial search engines and Scaffold accepts results from most. Common commercial search engines are Mascot and Sequest HT as part of Proteome Discoverer. Common open source search engines include MaxQuant, Comet and X! Tandem (run outside of Scaffold). These provide Peptide Spectrum Matches (PSMs). Please see our File Compatibility Matrix for more information on loading data into Scaffold.
Step 3. Load Data in Scaffold: Search engine output files are loaded into Scaffold using the file loading wizard. Scaffold accepts files in numerous different formats from commercial and open source search engines. While X! Tandem can be run in Scaffold raw data files cannot be loaded, X! Tandem within Scaffold only recognizes output files from a search engine. Once loaded, Scaffold provides the ability to view identified peptides and proteins, add GO annotations, apply statistical tests, and export Excel reports, mzIdentML files and unmatched spectra.