MultiNotch MS3 Enables Accurate, Sensitive, and Multiplexed Detection of Differential Expression across Cancer Cell Line Proteomes

By G. McAlister, David P. Nusinow, Mark P. Jedrychowski, M. Wühr, Edward L. Huttlin, B. Erickson, R. Rad, W. Haas, Steven P. Gygi. Analytical Chemistry, 2014

1,198 citations recorded by the source corpus.

Multiplexed quantitation via isobaric chemical tags (e.g., tandem mass tags (TMT) and isobaric tags for relative and absolute quantitation (iTRAQ)) has the potential to revolutionize quantitative proteomics. However, until recently the utility of these tags was questionable due to reporter ion ratio distortion resulting from fragmentation of coisolated interfering species. These interfering signals can be negated through additional gas-phase manipulations (e.g., MS/MS/MS (MS3) and proton-transfer reactions (PTR)). These methods, however, have a significant sensitivity penalty. Using isolation waveforms with multiple frequency notches (i.e., synchronous precursor selection, SPS), we coisolated and cofragmented multiple MS2 fragment ions, thereby increasing the number of reporter ions in the MS3 spectrum 10-fold over the standard MS3 method (i.e., MultiNotch MS3). By increasing the reporter ion signals, this method improves the dynamic range of reporter ion quantitation, reduces reporter ion signal variance, and ultimately produces more high-quality quantitative measurements. To demonstrate utility, we analyzed biological triplicates of eight colon cancer cell lines using the MultiNotch MS3 method. Across all the replicates we quantified 8 378 proteins in union and 6 168 proteins in common. Taking into account that each of these quantified proteins contains eight distinct cell-line measurements, this data set encompasses 174 704 quantitative ratios each measured in triplicate across the biological replicates. Herein, we demonstrate that the MultiNotch MS3 method uniquely combines multiplexing capacity with quantitative sensitivity and accuracy, drastically increasing the informational value obtainable from proteomic experiments.

Subfields: Molecular Biology, Spectroscopy

View Paper: https://www.semanticscholar.org/paper/d1d4c9284cabf410db306509c71cea9c0148fe20

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Citations in the corpus

Quantitative Proteomics of the Cancer Cell Line Encyclopedia (2020, positive): “MS3 quantification scans were performed using SPS selection (McAlister et al., 2014) of 10 notches from the MS2 spectrum and HCD fragmentation and readout in the Orbitrap at a 50k resolution and a maximum injection time of 150ms.”
Acute pharmacological degradation of Helios destabilizes regulatory T cells (2021, positive): “Each analysis used an MS3-based TMT method as described previously 39 .”
Multi-omics analysis identifies ATF4 as a key regulator of the mitochondrial stress response in mammals (2017, methodology): “, 2010), quantified by extracting the signal-to-noise ratio, and proteins were further collapsed to a final protein-level FDR of 1% (McAlister et al., 2014; Paulo et al., 2015).”
The MaxQuant computational platform for mass spectrometry-based shotgun proteomics (2016, methodology): “…distinguish between the MS level at which the quantification signals are generated, which can be in the MS1 spectra, as, for instance, with label-free or SILAC-based quantification; in the MS/MS spectra, as with conventional TMT labeling; or in the MS3 spectra, as with the multinotch approach 22 .”
Data‐independent acquisition‐based SWATH‐MS for quantitative proteomics: a tutorial (2018, positive): “…in reporter ions from co-eluting and co-fragmenting peptides, this has been addressed to some extent by using optimized data acquisition and analysis meth-ods (Ting et al , 2011; Savitski et al , 2013; McAlister et al , 2014; Ahrne et al , 2016; O’Brien et al , 2018; Sonnett et al , 2018).”
Obesity shapes metabolism in the tumor microenvironment to suppress anti-tumor immunity (2020, methodology): “Each analysis used an SPS-MS3-based TMT method (McAlister et al., 2014; Ting et al., 2011), which has been shown to reduce ion interference compared to MS2-based quantification (Paulo et al., 2016b).”
Autophagy promotes immune evasion of pancreatic cancer by degrading MHC-I (2020, positive): “All analysis involved an MS 3 -based TMT method as previously mentioned 51 and mass spectra was processed as described earlier 48 .”