Robust dimethyl-based multiplex-DIA doubles single-cell proteome depth via a reference channel
Publikation: Bidrag til tidsskrift › Tidsskriftartikel › Forskning › fagfællebedømt
Standard
Robust dimethyl-based multiplex-DIA doubles single-cell proteome depth via a reference channel. / Thielert, Marvin; Itang, Ericka C.M.; Ammar, Constantin; Rosenberger, Florian A.; Bludau, Isabell; Schweizer, Lisa; Nordmann, Thierry M.; Skowronek, Patricia; Wahle, Maria; Zeng, Wen Feng; Zhou, Xie Xuan; Brunner, Andreas David; Richter, Sabrina; Levesque, Mitchell P.; Theis, Fabian J.; Steger, Martin; Mann, Matthias.
I: Molecular Systems Biology, Bind 19, Nr. 9, e11503, 2023.Publikation: Bidrag til tidsskrift › Tidsskriftartikel › Forskning › fagfællebedømt
Harvard
APA
Vancouver
Author
Bibtex
}
RIS
TY - JOUR
T1 - Robust dimethyl-based multiplex-DIA doubles single-cell proteome depth via a reference channel
AU - Thielert, Marvin
AU - Itang, Ericka C.M.
AU - Ammar, Constantin
AU - Rosenberger, Florian A.
AU - Bludau, Isabell
AU - Schweizer, Lisa
AU - Nordmann, Thierry M.
AU - Skowronek, Patricia
AU - Wahle, Maria
AU - Zeng, Wen Feng
AU - Zhou, Xie Xuan
AU - Brunner, Andreas David
AU - Richter, Sabrina
AU - Levesque, Mitchell P.
AU - Theis, Fabian J.
AU - Steger, Martin
AU - Mann, Matthias
N1 - Publisher Copyright: © 2023 The Authors. Published under the terms of the CC BY 4.0 license.
PY - 2023
Y1 - 2023
N2 - Single-cell proteomics aims to characterize biological function and heterogeneity at the level of proteins in an unbiased manner. It is currently limited in proteomic depth, throughput, and robustness, which we address here by a streamlined multiplexed workflow using data-independent acquisition (mDIA). We demonstrate automated and complete dimethyl labeling of bulk or single-cell samples, without losing proteomic depth. Lys-N digestion enables five-plex quantification at MS1 and MS2 level. Because the multiplexed channels are quantitatively isolated from each other, mDIA accommodates a reference channel that does not interfere with the target channels. Our algorithm RefQuant takes advantage of this and confidently quantifies twice as many proteins per single cell compared to our previous work (Brunner et al, PMID 35226415), while our workflow currently allows routine analysis of 80 single cells per day. Finally, we combined mDIA with spatial proteomics to increase the throughput of Deep Visual Proteomics seven-fold for microdissection and four-fold for MS analysis. Applying this to primary cutaneous melanoma, we discovered proteomic signatures of cells within distinct tumor microenvironments, showcasing its potential for precision oncology.
AB - Single-cell proteomics aims to characterize biological function and heterogeneity at the level of proteins in an unbiased manner. It is currently limited in proteomic depth, throughput, and robustness, which we address here by a streamlined multiplexed workflow using data-independent acquisition (mDIA). We demonstrate automated and complete dimethyl labeling of bulk or single-cell samples, without losing proteomic depth. Lys-N digestion enables five-plex quantification at MS1 and MS2 level. Because the multiplexed channels are quantitatively isolated from each other, mDIA accommodates a reference channel that does not interfere with the target channels. Our algorithm RefQuant takes advantage of this and confidently quantifies twice as many proteins per single cell compared to our previous work (Brunner et al, PMID 35226415), while our workflow currently allows routine analysis of 80 single cells per day. Finally, we combined mDIA with spatial proteomics to increase the throughput of Deep Visual Proteomics seven-fold for microdissection and four-fold for MS analysis. Applying this to primary cutaneous melanoma, we discovered proteomic signatures of cells within distinct tumor microenvironments, showcasing its potential for precision oncology.
KW - DIA
KW - dimethyl labeling
KW - multiplexing
KW - single cells
KW - spatial proteomics
U2 - 10.15252/msb.202211503
DO - 10.15252/msb.202211503
M3 - Journal article
C2 - 37602975
AN - SCOPUS:85168570946
VL - 19
JO - Molecular Systems Biology
JF - Molecular Systems Biology
SN - 1744-4292
IS - 9
M1 - e11503
ER -
ID: 364546531