Sample sonication after trichloroacetic acid precipitation increases protein recovery from cultured hippocampal neurons, and improves resolution and reproducibility in two-dimensional gel electrophoresis.
Protein precipitation with TCA followed by acetone washing is frequently used to clean samples before 2-DE. However, the difficulty in solubilizing TCA-precipitated proteins causes some variability in 2-D gels and makes it difficult to detect some proteins. In this work we show that sonication of the samples, after TCA precipitation followed by elution in sample buffer, increases total protein recovery, and improves reproducibility and matching ratios between gels when analyzed by specialized software.
Manadas
Bruno J
BJ
Department of Zoology, Center for Neuroscience and Cell Biology, Dept. of Zoology, University of Coimbra, Coimbra, Portugal.
Vougas
Konstantinos
K
Fountoulakis
Michael
M
Duarte
Carlos B
CB
D000818:Animals; D002478:Cells, Cultured; D011232:Chemical Precipitation; D015180:Electrophoresis, Gel, Two-Dimensional / Q000379:methods*; D006624:Hippocampus / Q000737:chemistry* / Q000166:cytology; D009474:Neurons / Q000737:chemistry; D011506:Proteins / Q000737:chemistry / Q000302:isolation & purification*; D051381:Rats; D017208:Rats, Wistar; D015203:Reproducibility of Results; D013010:Sonication; D014238:Trichloroacetic Acid / Q000737:chemistry*
D011506:Proteins; D014238:Trichloroacetic Acid