Question:
Sample Clean Up: Why You Should Do It?
The Protein Man Says:
Protein sample clean up refers to the process of removing non-protein agents that can interfere with the results of downstream processes such as isoelectric focusing (IEF) and 2D electrophoresis. What can possibly happen if you don't clean up your protein samples prior to analysis? Here are some things you absolutely need to know.
Cleaning Up Your Protein Sample – Why Should You Do It?
Cleaning up your protein samples is considered to be of vital importance since the presence of interfering agents such as salts, ionic contaminants (lipids, ionic detergents and other phenolic compounds), nucleic acids, polysaccharides, plant-specific compounds (chlorophyll, lignin and tannins) and other charged agents may severely disrupt most electrophoresis techniques.
In addition, cleaning up your sample can also concentrate your protein solution to ensure better protein spot resolution and reduce spot streaking. By using an appropriate protein precipitation agent, you can transform your diluted protein samples (<1 ng/ml) into highly concentrated protein pellets that are virtually free from non-protein contaminants. These pellets can then be reconstituted in a small volume of the sample loading buffer prior to performing IEF and 2D electrophoresis studies to ensure more accurate results.
Contamination Removal Systems
There are a number of products that can be used to clean up your protein samples. These include the following:
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Dialysis systems – separates unwanted compounds from the macromolecules in your protein solution through selective diffusion
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Desalting and buffer exchange systems – desalting refers to the removal of salts from your protein sample while buffer exchange refers to the replacement of one set of buffer salts with another through size exclusion chromatography
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Detergent removal systems – you can remove detergents from your protein sample by using an appropriate precipitation system or a rapid column based system
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Electrophoresis clean-up systems – remove contaminants that can interfere with most electrophoretic analysis while increasing gel quality and reproducibility
Sample clean up can help you get more accurate results so make sure you don't skip this step before running any downstream processes needed in your protein research.
