Protein Identification by Peptide Mass Fingerprinting
Mass spectrometry has proven an important tool to the identification of proteins by the procedure call Peptide Mass Fingerprinting. In this process, targeted proteins are isolated by SDS gel electrophoresis and are digested directly in the gel slice with trypsin or Endo LysC C endoproteinase. The resulting peptides are extracted and the unfractionated mixture is analyzed by MALDI-TOF mass spectrometry. The mass measurements of the resulting peptides are used to query a database of protein and DNA sequences for likely candidate proteins. This is accomplished by matching the measured masses with masses predicted from the databases after 'virtual' digestion with the proteinase. The more peptide masses that match the predicted masses, the more certain one is of the likelihood that the protein is identified. Clearly, high-mass accuracy is required for this method to be of use. Sometimes no matches are found or the level of certainty is too low.
The success of a PMF protein identification experiment will depend upon reasonable sample estimates and very careful sample handling. We recommend that you contact us before planning any mass mapping experiments to help you estimate the amount of protein and describe preferred handling techniques. Experiments of this nature are also more successful when working with proteins from species with sequenced genomes. If you have any questions, please consult with the PCL staff before submitting a sample.