Comparison of different digestion methods for proteomic analysis of isolated cells and FFPE tissue samples
The result's identifiers
Result code in IS VaVaI
<a href="https://www.isvavai.cz/riv?ss=detail&h=RIV%2F00209805%3A_____%2F21%3A00078636" target="_blank" >RIV/00209805:_____/21:00078636 - isvavai.cz</a>
Result on the web
<a href="https://www.sciencedirect.com/science/article/pii/S0039914021004896" target="_blank" >https://www.sciencedirect.com/science/article/pii/S0039914021004896</a>
DOI - Digital Object Identifier
<a href="http://dx.doi.org/10.1016/j.talanta.2021.122568" target="_blank" >10.1016/j.talanta.2021.122568</a>
Alternative languages
Result language
angličtina
Original language name
Comparison of different digestion methods for proteomic analysis of isolated cells and FFPE tissue samples
Original language description
Proteomics of human tissues and isolated cellular subpopulations create new opportunities for therapy and monitoring of a patients' treatment in the clinic. Important considerations in such analysis include recovery of adequate amounts of protein for analysis and reproducibility in sample collection. In this study we compared several protocols for proteomic sample preparation: i) filter-aided sample preparation (FASP), ii) in-solution digestion (ISD) and iii) a pressure-assisted digestion (PCT) method. PCT method is known for already a decade [1], however it is not widely used in proteomic research. We assessed protocols for proteome profiling of isolated immune cell subsets and formalin-fixed paraffin embedded (FFPE) tissue samples. Our results show that the ISD method has very good efficiency of protein and peptide identification from the whole proteome, while the FASP method is particularly effective in identification of membrane proteins. Pressure-assisted digestion methods generally provide lower numbers of protein/peptide identifications, but have gained in popularity due to their shorter digestion time making them considerably faster than for ISD or FASP. Furthermore, PCT does not result in substantial sample loss when applied to samples of 50 000 cells. Analysis of FFPE tissues shows comparable results. ISD method similarly yields the highest number of identifications. Furthermore, proteins isolated from FFPE samples show a significant reduction of cleavages at lysine sites due to chemical modifications with formaldehyde-such as methylation (+14 Da) being among the most common. The data we present will be helpful for making decisions about the robust preparation of clinical samples for biomarker discovery and studies on pathomechanisms of various diseases.
Czech name
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Czech description
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Classification
Type
J<sub>imp</sub> - Article in a specialist periodical, which is included in the Web of Science database
CEP classification
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OECD FORD branch
10608 - Biochemistry and molecular biology
Result continuities
Project
<a href="/en/project/EF16_019%2F0000868" target="_blank" >EF16_019/0000868: Molecular, cellular and clinical approach to healthy ageing</a><br>
Continuities
P - Projekt vyzkumu a vyvoje financovany z verejnych zdroju (s odkazem do CEP)<br>I - Institucionalni podpora na dlouhodoby koncepcni rozvoj vyzkumne organizace
Others
Publication year
2021
Confidentiality
S - Úplné a pravdivé údaje o projektu nepodléhají ochraně podle zvláštních právních předpisů
Data specific for result type
Name of the periodical
Talanta
ISSN
0039-9140
e-ISSN
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Volume of the periodical
233
Issue of the periodical within the volume
october 2021
Country of publishing house
NL - THE KINGDOM OF THE NETHERLANDS
Number of pages
11
Pages from-to
122568
UT code for WoS article
000668000500083
EID of the result in the Scopus database
2-s2.0-85108280622