MassIVE MSV000090624

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IP-LC/MS pulldown assay for identifying TFPI2-associated membrane protein in human primary microglia

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Description

IP-LC/MS pulldown assay for identifying TFPI2-associated membrane protein in human primary microglia. Tandem mass spectra were extracted. Charge state deconvolution and deisotoping were not performed. All MS/MS samples were analyzed using Mascot (Matrix Science, London, UK; version 2.8.0). Mascot was set up to search the uniprot-SP-human_20190906_20200629 database (20432 entries) assuming the digestion enzyme trypsin. Mascot was searched with a fragment ion mass tolerance of 0.050 Da and a parent ion tolerance of 10.0 PPM. Carbamidomethyl of cysteine was specified in Mascot as a fixed modification. Deamidated of asparagine and glutamine, oxidation of methionine and acetyl of the n-terminus were specified in Mascot as variable modifications. Scaffold (version Scaffold_5.1.2, Proteome Software Inc., Portland, OR) was used to validate MS/MS based peptide and protein identifications. Peptide identifications were accepted if they could be established at greater than 90.0% probability by the Peptide Prophet algorithm (Keller, A et al Anal. Chem. 2002;74(20):5383-92) with Scaffold delta-mass correction. Protein identifications were accepted if they could be established at greater than 99.0% probability to achieve an FDR less than 1.0% and contained at least 2 identified peptides. Protein probabilities were assigned by the Protein Prophet algorithm (Nesvizhskii, Al et al Anal. Chem. 2003;75(17):4646-58). Proteins that contained similar peptides and could not be differentiated based on MS/MS analysis alone were grouped to satisfy the principles of parsimony. Sample #3 is protein eluted from the complex containing TFPI2 antibody. Sample #4 is IgG control. [doi:10.25345/C5CR5NH6W] [dataset license: CC0 1.0 Universal (CC0 1.0)]

Keywords: TFPI2, Microglia, Glioblastoma

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Principal Investigators:
(in alphabetical order) 		Peiwen Chen, Northwestern University, US
Submitting User: Lizhi
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