Robustness and Ruggedness of Isoelectric Focusing and Superficially Porous Liquid Chromatography with Fourier Transform Mass Spectrometry.

ABSTRACT: An investigation of a multidimensional proteomics workflow composed of off-gel isoelectric focusing (IEF) and superficially porous liquid chromatography (SPLC) with Fourier transform mass spectrometry (FTMS) was completed in order to assess various figures of merit associated with intact protein measurements. Triplicate analysis performed at both high and low FTMS resolutions on the E. coli proteome resulted in ∼900 redundant proteoforms from 3 to 95 kDa. Normalization of the chromatographic axis to identified proteoforms enabled reproducible physicochemical property measurements between proteome replicates with inter-replicate variances of ±3 ppm mass error for proteoforms <30 kDa, ±1.1 Da for proteins >30 kDa, ±12 s retention time error, and ±0.21 pI units. The results for

SUBMITTER: Corbett JR 

PROVIDER: S-EPMC10476448 | biostudies-literature | 2021 Jan

REPOSITORIES: biostudies-literature