Protein quantitation workflows using the TripleTOF 6600: a case study for Rituximab (SCIEX)
Although the triple-stage quadrupole (QQQ) mass spectrometer remains the pillar for quantitative LC-MS/MS bioanalytical assays, due in part to the platform’s high duty cycle when operated in multiple-reaction monitoring (MRM) mode, the applicability of highresolution mass spectrometry (HRMS) has become of increasing importance for protein quantitation given the complexity of proteolytically digested samples in the surrogate peptide approach. While the QQQ demonstrates high sensitivity and specificity, the relatively low-resolution measurement of m/z may fail to differentiate analyte response from nominally isobaric background interference.
In contrast, HRMS with accurate mass assignment of product ion allows interference to be resolved through judicious selection of a post-acquisition mass extraction window whose tolerance is largely dictated by the effective resolution and stability of mass calibration.
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