ECD is a great tool  for the analysis of very labile or otherwise difficult modifications. Typical examples for these are

  • phosphorylation (in particular on Lys, Arg and His residues)
  • deamidation (Asp/isoAsp formation which cannot be differentiated by collision induced dissociation)
  • sulfatation
  • γ-carboxyglutamate formation
  • modifications where the context on the intact protein is relevant such as deciphering the histone code

The reason why ECD is particularly suited is due to the nature of the activation. Vibrational activation by CID tends to break the weakest bonds first which leads to losses of labile modifications. In contrast to CID, the electronic activation by ECD leaves these labile bonds intact and allows for the identification and localization of the modified residues. Due to ECD’s top-down capabilities, also the complex task of deciphering multiple modifications in their context, as it is a typical task in histone analysis, is possible.

A common problem in bioplogics analysis is also protein aging. A frequent observation in this case is the deamidation of Asparagine residues resulting in a mixture of Aspartic acid and isoAspartic acid. As these two residues are identical in mass and share the same bonds with their neighbouring residues, they cannot be differentiated by CID. In contrast, ECD enables you to directly identify the presence of isoAspartic acid by formation of a characteristic z-57 fragment for isoAsp. In most cases this overcomes the time consuming need to generate reference peptide for analysis by LC-MS against the reference standards.