High accuracy mass measurement of peptides with internal calibration using a dual electrospray ionization sprayer...

A dual-ESI-sprayer system was constructed and applied to achieve high accuracy of peptide mass measurement for protein identification by means of peptide mapping. Sample was introduced in one sprayer, and reference in the other, thus making internal calibration possible greatly enhancing the mass accuracy. Several samples were utilized to evaluate the reliability of this dual-ESI-sprayer system. The range of mass errors was 0.16-5.37 ppm. The peptide masses of tryptic digests of myoglobin (horse) were measured by the HPLC/dual-ESI-MS system, with mass deviations ranging from 0.01-7.67 ppm, and about 75% mass deviations below 5 ppm with 40% below 1[?]ppm. These peptide masses were utilized to perform database searching for protein identification, and compared to results obtained by external calibration. This comparison showed that the internal calibration provides a more reliable method of protein identification, with a much smaller number of required peptides for matching, and with less CPU time consumed for database searching.