Improvement in Peptide Detection for Proteomics Analyses Using NanoLC−MS and High-Field Asymmetry Waveform Ion Mobility Mass Spectrometry

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Abstract
Sensitive and selective detection of multiply charged peptide ions from complex tryptic digests was achieved using high-field asymmetric waveform ion mobility spectrometry (FAIMS) combined with nanoscale liquid chromatography−mass spectrometry (nanoLC−FAIMS-MS). The combination of FAIMS provided a marked advantage over conventional nanoLC−MS experiments by reducing the extent of chemical noise associated with singly charged ions and enhancing the overall population of detectable tryptic peptides. Such advantages were evidenced by a 6−12-fold improvement in signal-to-noise ratio measurements for a wide range of multiply charged peptide ions. An increase of 20% in the number of detected peptides compared to conventional nanoelectrospray was achieved by transmitting ions of different mobilities at high electric field vs low field while simultaneously recording each ion population in separate mass spectrometry acquisition channels. This method provided excellent reproducibility across replicate nanoLC−FAIMS-MS runs with more than 90% of all detected peptide ions showing less than 30% variation in intensity. The application of this technique in the context of proteomics research is demonstrated for the identification of trace-level proteins showing differential expression in U937 monocyte cell extracts following incubation with phorbol ester.