Issue Information. (24th September 2020)

• Record Type: Journal Article
• Is Part Of: Current protocols in immunology. Volume 130(2020)
• Page Start: n/a
• Page End: n/a
• Publication Date: 2020-09-24
• Subjects: Immunology -- Laboratory manuals; Medical protocols; Immunologic Techniques; Medical protocols; Immunology; Laboratory manuals; Reference works; 571.96028
• DOI: 10.1002/cpim.81
• Languages: English
• ISSNs: 1934-3671

Abstract

Abstract : Cover : In Brian et al. (http://doi.org/10.1002/cpim.104 ), the image shows key steps for in‐gel digestion and targeted LC‐MS/MS. Immunoprecipitated samples are resolved via gel electrophoresis and excised based on molecular weight. Protein standards (e.g., bovine serum albumin [BSA]) of known concentration are used to quantify gel loading to standardize loading of immunoprecipitate samples onto the LC‐MS/MS and normalizing later peptide or phosphopeptide analysis. A reference quantity of isotope‐labeled reference peptide, to be used as an internal standard, is added to the gel fragments. The gel sample/peptide mixture is subjected to protease digest and targeted LC‐MS/MS analysis. By comparing the endogenous and isotope‐labeled phosphopeptide peaks and peptide standard curves, molar and relative quantities of phosphorylated and unphosphorylated peptides can be calculated.