Peptide mass mapping data analysis

The introduction of delayed ion extraction in MALDI-TOF instruments in 1995 resulted in a tremendous improvement in analytical performance, mass resolution and accuracy. Using this technique, peptide mass mapping gained high specificity for protein identification and an ability to identify individual components among simple protein mixtures [60, 61]. Improved sample preparation methods also provided an enhanced sensitivity to enable analysis of femtomolar levels of protein [5, 19], Today, the automation of MALDI-MS data acquisition enables the analysis of hundreds of samples each day in large-scale protein identification experiments [62]. 

Matrix-assisted laser desorption-ionization ionizes molecules with molecular masses of 100-1,000,000 Da for analysis by MS and provides high sensitivity, high throughput, and simplicity of operation. MALDI combined with enzymatic reactions and protein chemistry can provide very useful information on molecular masses, peptide maps, and primary structure.15 MALDI-TOF MS can quickly and accurately determine unfractionated mixtures at concentrations below 100 fmol per liter. The obtained data are then calibrated with internal standards, monoisotopic masses are assigned for all prominent peaks, and the peptide list thus generated is used to identify the protein by using a suitable database.16... 

Traditional methods to generate peptide maps involve fractionation of complex mixtures of peptides in a protein digest either with one-dimensional SDS-PAGE or RP-HPLC [28,29]. The mass spectrometry peptide-mapping protocol, in principle, is similar to these techniques, but it provides an added dimension of structure-specific data (i.e., the molecular mass). MALDI-MS [30,31], ESl-MS [32], LC/ESI-MS [33], and CE/ESI-MS [34] have currently replaced the traditional biochemical approaches. MALDI allows the direct analysis of unfractionated protein digests. The commonly used matrices are sinapinic acid, a-cyano-4-hydroxy cinnamic acid (a-CHCA), and 2,5-dihydroxybenzoic acid (DHB). 

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