MassMatrix is a program that matches tandem mass spectra with theoretical peptide sequences derived from a protein database. The program uses a mass accuracy sensitive probabilistic score model to rank peptide matches. The MS/MS search software was evaluated by use of a high mass accuracy dataset and its results compared with those from MASCOT, SEQUEST, X!Tandem, and OMSSA. For the high mass accuracy data, MassMatrix provided better sensitivity than MASCOT, SEQUEST, X!Tandem, and OMSSA for a given specificity and the percentage of false positives was 2%. More importantly all manually validated true positives corresponded to a unique peptide/spectrum match. The presence of decoy sequence and additional variable PTMs did not significantly affect the results from the high mass accuracy search. MassMatrix performs well when compared with MASCOT, SEQUEST, X!Tandem, and OMSSA with regard to search time. MassMatrix was also run on a distributed memory clusters and achieved search speeds of approximately 100,000 spectra per hour when searching against a complete human database with eight variable modifications. The algorithm is available for public searches at (http://www.massmatrix.net).