EVs were lysed in the radioimmunoprecipitation assay (RIPA) buffer and total proteins were digested using the S‐TRAP Micro column. Each sample was loaded on nanoEase M/Z C18 5‐μm trap column, eluted on C18 1.7‐μm column (Waters, USA) and analyzed using LC‐MS/MS on the TripleTOF 6600 system (Sciex, Canada). Peptide identification was performed with the ProteinPilot 5.0.3 software (Sciex, Canada) against the SwissProt Homo sapiens database. The relative abundance of each identified protein was determined using the Mascot Server software (Matrix Science, UK). This analysis was performed by the Mass Spectrometry facility in the Protein and Proteomics Centre, National University of Singapore, Singapore. Peptides with confidence level ≥95% were used for further analysis. Gene Ontology (GO) enrichment of biological process, molecular function, and cellular component of identified proteins was performed using g:Profiler.
Mascot server software
Manufactured by Matrix Science
Sourced in United Kingdom, United States
Mascot Server software is a search engine used for protein identification and characterization from mass spectrometry data. It provides a platform for automated, high-throughput analysis of proteomic samples.
Automatically generated - may contain errors
2 protocols using mascot server software
1
EV Protein Identification and Analysis
2
Identification of Phaseolus vulgaris Proteins
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Protein (25 µg) was loaded on SDS-PAGE and bands corresponding to the non-processed (approximately 35 kDa) and processed forms (a combination of α- and β- subunits of αAI, with a molecular mass between 15 and 20 kDa) were excised from the gel. Samples were submitted to Australian Proteome Analysis Facility (Macquarie University, Sydney, NSW, Australia), digested with trypsin and loaded onto a capillary LC system coupled to an MS/MS instrument (1D Nano LC ESI MS/MS) as described by Atack et al. [42 (link)]. Tryptic fragments were identified using Mascot Server software (Matrix Science Inc., MA, USA) using the Phaseolus_vulgaris database.
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