Protein sample preparation workflow.
Due to the great diversity of protein sample types and sources, no single sample preparation method works with all proteins; for any sample, the optimum procedure must be determined empirically. However, the following general sample preparation guidelines should be kept in mind to avoid a number of pitfalls during sample preparation for protein electrophoresis (Posch et al. 2006).
- Keep the sample preparation workflow as simple as possible (increasing the number of sample handling steps may increase variability)
- With cell or tissue lysates, include protease inhibitors to minimize artifacts generated by proteolysis; protease inhibitors are generally not required for samples like serum or plasma
- Solubilize proteins in a buffer that is compatible with the corresponding electrophoresis technique
- Determine the amount of total protein in each sample using a protein assay that is compatible with chemicals in your samples
- Use protein extracts immediately or aliquot them into appropriately sized batches and store them at –70°C to avoid freeze-thaw cycles
Posch A et al. (2006). Tools for sample preparation and prefractionation in two-dimensional gel electrophoresis. In Separation methods in Proteomics, Smejkal GB ed. (Boca Raton: CRC Press), pp 107–133.