Comparison of sodium dodecyl sulfate depletion techniques for proteome analysis by mass spectrometry
Comparison of sodium dodecyl sulfate depletion techniques for proteome analysis by mass spectrometry
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DOI：
10.1016/j.chroma.2015.09.042
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发表时间：
2015-10-30
影响因子：
4.1
通讯作者：
Doucette, Alan
中科院分区：
文献类型：
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作者：
Kachuk, Carolyn;Stephen, Kegan;Doucette, Alan
在蛋白质组学中，十二烷基硫酸钠（SDS）有利于蛋白质溶解和基于质量的分离（例如GELFrEE或SOS PAGE）。许多SDS耗尽技术可用于在质谱法之前纯化蛋白质。纯化的有效性对分析的成功具有控制性影响。在这里，我们定量评估了SDS消耗的八种方法：凝胶内消化;在丙酮中或用TCA沉淀蛋白质;用KCl沉淀去污剂;强阳离子交换;用Pierce去污剂去除柱进行蛋白质水平和肽水平纯化;和FASP II。考虑到蛋白质纯度，FASP II显示出最高程度的SOS去除，与凝胶内消化相匹配（超过99.99%耗尽）。其他方法（丙酮、强阳离子交换、Pierce柱）也将SOS消耗至适合LC-MS的水平（>99%）。考虑到蛋白质回收率，FASP II显示了显著的样品损失（
In proteomics, sodium dodecyl sulfate (SDS) is favored for protein solubilization and mass-based separation (e.g. GELFrEE or SOS PAGE). Numerous SDS depletion techniques are available to purify proteins ahead of mass spectrometry. The effectiveness of the purification has a controlling influence on the success of the analysis. Here we quantitatively assess eight approaches to SDS depletion: in-gel digestion; protein precipitation in acetone or with TCA; detergent precipitation with KCI; strong cation exchange; protein level and peptide level purification with Pierce detergent removal cartridges; and FASP II. Considering protein purity, FASP II showed the highest degree of SOS removal, matching that of in-gel digestion (over 99.99% depleted). Other methods (acetone, strong cation exchange, Pierce cartridges) also deplete SOS to levels amenable to LC-MS (>99%). Accounting for protein recovery, FASP II revealed significant sample loss (