SIMSI-Transfer: Software-Assisted Reduction of Missing Values in Phosphoproteomic and Proteomic Isobaric Labeling Data Using Tandem Mass Spectrum Clustering

Hamood, Firas; Bayer, Florian; Wilhelm, Mathias; Küster, Bernhard; The, Matthew

doi:10.1016/j.mcpro.2022.100238

Cited by 11 publications

(10 citation statements)

References 34 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…The solution would be the further improvement in instruments and data analysis tools. Very recently, SIMSI-transfer was developed to transfer successful database search hits in one TMT batch to other TMT batches to help rescue imperfect MS2 spectra based on the similarity of MS2 spectra and the accurate mass, by which the proteome coverage in each batch and the overlap percentage among all batches are remarkably increased . Similar algorithms could be developed for TAG-TMTpro approach to increase the proteome coverage and the identification overlap.…”

Section: Discussionmentioning

confidence: 99%

TAG-TMTpro, a Hyperplexing Quantitative Approach for High-Throughput Proteomic Studies

Shen

Zhang

2022

Anal. Chem.

View full text Add to dashboard Cite

Isobaric labeling is the most widely used multiplexing quantitative approach in proteomic studies, enabling the comparison of up to 18 samples in a single MS analysis. Expanding the multiplexing capacity is of great necessity for high-throughput proteomic studies. Herein, we establish a novel TAG-TMTpro approach by introducing Ala or Gly residues to peptides prior to TMTpro labeling, which is able to triple the quantitative capacity of TMTpro. We systematically evaluated the Boc-Ala-OSu and Boc-Gly-OSu reaction and optimized the conditions for labeling, side-product elimination, and Boc deprotection. We validated the identification and quantification performance using E. coli and HeLa cell lysates. We demonstrated that the TAG-TMTpro approach resulted in good identification reproducibility and reliable quantitative accuracy. The TAG-TMTpro is able to triple the multiplexing capacity of TMTpro reagents and is a versatile quantitative approach for high-throughput proteomic studies. Data are available via ProteomeXchange with identifier PXD033711.

show abstract

Section: Discussionmentioning

confidence: 99%

TAG-TMTpro, a Hyperplexing Quantitative Approach for High-Throughput Proteomic Studies

Shen

Zhang

2022

Anal. Chem.

View full text Add to dashboard Cite

show abstract

“…Currently, we used a 50× carrier to obtain good proteome coverage in the RTS-based protein identification and quantification; however, we suggest that the carrier-sample ratio should be further reduced to 10× or lower with the sensitivity improvement of the RTS algorithm. The reduced proteome coverage due to low carrier ratio could be improved using isobaric match between runs (iMBR) algorithm, as demonstrated by recent studies. , In addition to sensitivity and quantification accuracy, other technical improvements such as analysis throughput could be achieved by using higher multiplex isobaric reagents or combined precursor isotopic labeling and isobaric tagging (cPILOT) approaches. , Moreover, because of the inherent differences in ion detection between LIT and OT, it could be highly valuable to perform an additional study on the response linearity and quantification dynamic ranges of both detection methods for single-cell-scale samples. Such study could provide insight on their performance in the absolute quantification of proteins.…”

Section: Discussionmentioning

confidence: 99%

“…The reduced proteome coverage due to low carrier ratio could be improved using isobaric match between runs (iMBR) algorithm, as demonstrated by recent studies. 65,66 In addition to sensitivity and quantification accuracy, other technical improvements such as analysis throughput could be achieved by using higher multiplex isobaric reagents 67 or combined precursor isotopic labeling and isobaric tagging (cPILOT) approaches. 68,69 Moreover, because of the inherent differences in ion detection between LIT and OT, it could be highly valuable to perform an additional study on the response linearity and quantification dynamic ranges of both detection methods for single-cell-scale samples.…”

Section: ■ Conclusionmentioning

confidence: 99%

Evaluating Linear Ion Trap for MS3-Based Multiplexed Single-Cell Proteomics

Park

Fulcher

et al. 2023

Anal. Chem.

View full text Add to dashboard Cite

There is a growing demand to develop high-throughput and high-sensitivity mass spectrometry methods for single-cell proteomics. The commonly used isobaric labeling-based multiplexed single-cell proteomics approach suffers from distorted protein quantification due to co-isolated interfering ions during MS/MS fragmentation, also known as ratio compression. We reasoned that the use of MS3-based quantification could mitigate ratio compression and provide better quantification. However, previous studies indicated reduced proteome coverages in the MS3 method, likely due to long duty cycle time and ion losses during multilevel ion selection and fragmentation. Herein, we described an improved MS acquisition method for MS3-based single-cell proteomics by employing a linear ion trap to measure reporter ions. We demonstrated that linear ion trap can increase the proteome coverages for single-cell-level peptides with even higher gain obtained via the MS3 method. The optimized real-time search MS3 method was further applied to study the immune activation of single macrophages. Among a total of 126 single cells studied, over 1200 and 1000 proteins were quantifiable when at least 50 and 75% nonmissing data were required, respectively. Our evaluation also revealed several limitations of the low-resolution ion trap detector for multiplexed single-cell proteomics and suggested experimental solutions to minimize their impacts on single-cell analysis.

show abstract

“…The labeling method with TMT detected changes in the number of low-abundance proteins available for hypothesis testing, showing higher precision and fewer missing values compared to a label-free quantitation method [ 42 ]. It should be noted that the accurate interpretation of TMT-based quantitative proteomic data requires minimizing false positives, batch effects, and missing values [ 43 , 44 ].…”

Section: Tmt-assisted Single-cell Proteomicsmentioning

confidence: 99%

Advances in Mass Spectrometry-Based Single Cell Analysis

Lee

et al. 2023

Biology

View full text Add to dashboard Cite

Technological developments and improvements in single-cell isolation and analytical platforms allow for advanced molecular profiling at the single-cell level, which reveals cell-to-cell variation within the admixture cells in complex biological or clinical systems. This helps to understand the cellular heterogeneity of normal or diseased tissues and organs. However, most studies focused on the analysis of nucleic acids (e.g., DNA and RNA) and mass spectrometry (MS)-based analysis for proteins and metabolites of a single cell lagged until recently. Undoubtedly, MS-based single-cell analysis will provide a deeper insight into cellular mechanisms related to health and disease. This review summarizes recent advances in MS-based single-cell analysis methods and their applications in biology and medicine.

show abstract

SIMSI-Transfer: Software-Assisted Reduction of Missing Values in Phosphoproteomic and Proteomic Isobaric Labeling Data Using Tandem Mass Spectrum Clustering

Cited by 11 publications

References 34 publications

TAG-TMTpro, a Hyperplexing Quantitative Approach for High-Throughput Proteomic Studies

TAG-TMTpro, a Hyperplexing Quantitative Approach for High-Throughput Proteomic Studies

Evaluating Linear Ion Trap for MS3-Based Multiplexed Single-Cell Proteomics

Advances in Mass Spectrometry-Based Single Cell Analysis

Contact Info

Product

Resources

About