Enhanced Diagnostic Tools
LC-MS/MS: Nano HPLC coupled to an ion-trap or a QqTOF mass spectrometer. Analysis of proteolytic fragments.
MALDI-TOF Based Peptide Mass Mapping: Mass determination of proteolytic fragments.
Edman Seqeuncing: Chemical sequence determination of intact, unblocked proteins or proteolytic fragments
In general, the LC-MS/MS method will yield the highest confidence identifications and is the method of choice, provided the expected protein sequence or a highly homologous sequence is in the searched database. If you are dealing with an organism with a relatively small genome, MALDI-TOF peptide mass mapping analysis may be sufficient for your needs. If you are dealing with an organism whose genome is not known or you wish to obtain N-terminal sequence information on an intact protein, Edman sequencing analysis is probably your best choice since it yields sequence information.
Both Edman sequencing and LC-MS/MS are useful for looking at protein modifications such as phosphorylations, acetylations, methylations, etc. However, due to the relatively low resolution characteristics of our ion-trap instrument, unambiguous assignments of modification sites can be difficult with this instrument. Edman sequencing can offer more unambiguous identification of modification sites, provided that suitable standards for the modified amino acids are available. Edman sequencing also suffers from the limitations listed above.
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