Tandem MS Peptide Sequencing

The experimental MS/MS spectra are matched against theoretical spectra and cross correlation scores are calculated based on the extent to which the predicted and experimental spectra overlap.8 The higher cross correlation scores reflect a high level of matching of the experimental and predicted MS/MS spectra, and vice versa. The difference between a normalized cross correlation score and the next best match is reported as the (ACn) and indicates the quality of the top match in comparison to the next ranked sequences in the database.8 

Several algorithms are available for the analysis of MS/MS spectra including SEQUEST, MASCOT, and X Tandem among others. Note that additional secondary quality control of assessment of MS/MS data has recently been implemented to assess identification probabilities and false positivity rates. The MS/MS spectra from an experiment can be interrogated against a concatenated forward and reverse database and an assessment of the intrinsic error rate of the data set can be made. Other approaches for secondary analysis of matching scores for peptide sequencing data include XCorr score normalization routines that are independent of peptide and database size.33 

Tandem MS has emerged as a definitive approach for identifying proteins from multiple sources including complex mixtures. In comparison to PMF, MS/MS permits more definitive identification of proteins. Matching of multiple MS/MS spectra to peptide sequences within the same protein 

---