Journal
PROTEOMICS
Volume 16, Issue 13, Pages 1847-1851Publisher
WILEY-BLACKWELL
DOI: 10.1002/pmic.201600024
Keywords
Acetone; Protein precipitation; SWATH; TCA/acetone; Technology
Funding
- Fundacao para a Ciencia e Tecnologia (FCT) [PTDC/SAU-NEU/103728/2008, PTDC/NEU-NMC/0205/2012, UID/NEU/04539/2013]
- COMPETE Programa Operacional Factores de Competitividade, QREN
- European Union (FEDER - Fundo Europeu de Desenvolvimento Regional)
- National Mass Spectrometry Network (RNEM) [REDE/1506/REM/2005]
- FCT [SFRH/BD/88419/2012, SFRH/BD/81495/2011]
- Fundação para a Ciência e a Tecnologia [PTDC/SAU-NEU/103728/2008, SFRH/BD/88419/2012] Funding Source: FCT
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Proteomic approaches are extremely valuable in many fields of research, where mass spectrometry methods have gained an increasing interest, especially because of the ability to perform quantitative analysis. Nonetheless, sample preparation prior to mass spectrometry analysis is of the utmost importance. In this work, two protein precipitation approaches, widely used for cleaning and concentrating protein samples, were tested and compared in very diluted samples solubilized in a strong buffer (containing SDS). The amount of protein recovered after acetone and TCA/acetone precipitation was assessed, as well as the protein identification and relative quantification by SWATH-MS yields were compared with the results from the same sample without precipitation. From this study, it was possible to conclude that in the case of diluted samples in denaturing buffers, the use of cold acetone as precipitation protocol is more favourable than the use of TCA/acetone in terms of reproducibility in protein recovery and number of identified and quantified proteins. Furthermore, the reproducibility in relative quantification of the proteins is even higher in samples precipitated with acetone compared with the original sample.
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