Searching With Peptide Mass Fingerprints.
In this method, first a mixture of protein is subjected to 2nd gel electrophoresis. A specific protein is then used for in gel digestion with a sequence-specific protease such as trypsin and a mass spectrum described as mass fingerprint is obtained by MALDI-TOF. The mass data are compared to the theoretically expected tryptic peptide masses for each entry in the database. The proteins in the databases may differ according to the number of peptide matches.

More sophisticated scoring algorithms take the mass accuracy and the proportion of protein covered into account and attempts to calculate a level of confidence for the match. When high mass accuracy is needed, as a rule at least five peptide masses need to be matched to the protein and 15% of the protein sequence needs to be covered for an unambiguous identification. After a match has been found, a second-pass search is performed to correlate peptides with the database sequence of the match, taking into account possible modification.