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Matrix‐assisted laser desorption/ionization‐MS‐based relative quantification of peptides and proteins using iodoacetamide and N ‐methyliodoacetamide as labeling reagents
Author(s) -
Sun MeiChuan,
Chen ChengDong,
Huang YuShan,
Wu ZhongSiang,
Ho YenPeng
Publication year - 2008
Publication title -
journal of separation science
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.72
H-Index - 102
eISSN - 1615-9314
pISSN - 1615-9306
DOI - 10.1002/jssc.200700440
Subject(s) - iodoacetamide , chemistry , chromatography , reagent , lysozyme , matrix assisted laser desorption/ionization , cysteine , alkylation , isobaric labeling , denaturation (fissile materials) , matrix (chemical analysis) , mass spectrometry , desorption , tandem mass spectrometry , biochemistry , enzyme , organic chemistry , protein mass spectrometry , adsorption , nuclear chemistry , catalysis
The use of iodoacetamide (IAA) and N ‐methyliodoacetamide (MIAA) as labeling agents for the relative measurements of proteins using MALDI‐MS is described herein. These reagents, which alkylate the thiol groups of cysteine residues in proteins, were introduced during the alkylation step of a common protein denaturation and digestion process. This approach is simpler and cheaper than those involving isotope labeling agents. The labeling agents described herein displayed good dynamic ranges and correlation coefficients for protein quantification analyses when the proteins were treated through either in‐solution or in‐gel digestion. The best dynamic ranges (in the molar ratio) for proteins lysozyme, transferrin, and BSA (in‐solution digestion) are 0.1–10, 0.1–8, and 0.1–8, respectively. The corresponding correlation coefficients are greater than 0.99. The IAA/MIAA labeling is a useful method for the relative quantification of peptides and digested proteins when the chromatographic isotope effect is not a major concern.