Pre-extraction sample handling by automated frozen disruption significantly improves subsequent proteomic analyses

被引:40
作者
Butt, RH
Coorssen, JR
机构
[1] Univ Calgary, Fac Med, Hotchkiss Brain Inst, Calgary, AB, Canada
[2] Univ Calgary, Fac Med, Dept Physiol & Biophys, Calgary, AB, Canada
[3] Univ Calgary, Fac Med, Dept Biochem & Mol Biol, Calgary, AB, Canada
[4] Univ Calgary, Fac Med, Dept Cell Biol & Anat, Calgary, AB, Canada
关键词
proteomics; membrane proteins; tissue homogenization; protein extraction;
D O I
10.1021/pr0503634
中图分类号
Q5 [生物化学];
学科分类号
071010 ; 081704 ;
摘要
Here we quantitatively characterize two common homogenization strategies in the analysis of tissue proteomes: classical manual homogenization (MH) and an automated frozen disruption (AFD) technique. In a variety of tissues, many proteins were more efficiently extracted, resolved and detected, with high reproducibility after AFD, amounting to as much as 2% of the total resolved proteome. The benefits of AFD over MH are 2-fold: (1) AFD yields a much more thorough homogenate than MH; and (2) as a deep frozen alternative, AFD maintains a level of biological complexity that is not retained during MH. Thus, AFD coupled with refined 2DE protocols and Sypro Ruby staining yields quantitative proteomic analyses.
引用
收藏
页码:437 / 448
页数:12
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