SP3 Protocol for Proteomic Plant Sample Preparation Prior LC-MS/MS

@article{1768079,
   author = {Mikulášek, Kamil and Konečná, Hana and Potěšil, David and Holánková, Renata and Havliš, Jan and Zdráhal, Zbyněk},
   article_number = {MAR},
   doi = {http://dx.doi.org/10.3389/fpls.2021.635550},
   keywords = {bottom-up; protein cleanup; Arabidopsis thaliana; sodium dodecyl sulfate removal; single-pot solid-phase-enhanced sample preparation; carboxylated magnetic beads; filter-aided sample preparation; mass spectrometry},
   language = {eng},
   issn = {1664-462X},
   journal = {Frontiers in Plant Science},
   title = {SP3 Protocol for Proteomic Plant Sample Preparation Prior LC-MS/MS},
   url = {https://www.frontiersin.org/articles/10.3389/fpls.2021.635550/full},
   volume = {12},
   year = {2021}
}

TY  - JOUR
ID  - 1768079
AU  - Mikulášek, Kamil - Konečná, Hana - Potěšil, David - Holánková, Renata - Havliš, Jan - Zdráhal, Zbyněk
PY  - 2021
TI  - SP3 Protocol for Proteomic Plant Sample Preparation Prior LC-MS/MS
JF  - Frontiers in Plant Science
VL  - 12
IS  - MAR
SP  - "635550"
EP  - "635550"
PB  - Frontiers Research Foundation
SN  - 1664462X
KW  - bottom-up
KW  - protein cleanup
KW  - Arabidopsis thaliana
KW  - sodium dodecyl sulfate removal
KW  - single-pot solid-phase-enhanced sample preparation
KW  - carboxylated magnetic beads
KW  - filter-aided sample preparation
KW  - mass spectrometry
UR  - https://www.frontiersin.org/articles/10.3389/fpls.2021.635550/full
N2  - Quantitative protein extraction from biological samples, as well as contaminants removal before LC-MS/MS, is fundamental for the successful bottom-up proteomic analysis. Four sample preparation methods, including the filter-aided sample preparation (FASP), two single-pot solid-phase-enhanced sample preparations (SP3) on carboxylated or HILIC paramagnetic beads, and protein suspension trapping method (S-Trap) were evaluated for SDS removal and protein digestion from Arabidopsis thaliana (AT) lysate. Finally, the optimized carboxylated SP3 workflow was benchmarked closely against the routine FASP. Ultimately, LC-MS/MS analyses revealed that regarding the number of identifications, number of missed cleavages, proteome coverage, repeatability, reduction of handling time, and cost per assay, the SP3 on carboxylated magnetic particles proved to be the best alternative for SDS and other contaminants removal from plant sample lysate. A robust and efficient 2-h SP3 protocol for a wide range of protein input is presented, benefiting from no need to adjust the amount of beads, binding and rinsing conditions, or digestion parameters.
ER  - 

MIKULÁŠEK, Kamil, Hana KONEČNÁ, David POTĚŠIL, Renata HOLÁNKOVÁ, Jan HAVLIŠ and Zbyněk ZDRÁHAL. SP3 Protocol for Proteomic Plant Sample Preparation Prior LC-MS/MS. \textit{Frontiers in Plant Science}. Frontiers Research Foundation, 2021, vol.~12, MAR, p.~''635550'', 12 pp. ISSN~1664-462X. Available from: https://dx.doi.org/10.3389/fpls.2021.635550.