Protein sample preparation generally involves some or all of the following steps: cell disruption, extraction, removal of interfering compounds, changes of the physio/chemical properties and concentration of the sample. The quality of sample preparation can greatly affect the quality of the data that are generated. General guidelines and some of the most common methods for protein sample preparation are provided in this section. Protocols for sample preparation from mammalian tissue, plant leaves, microbial cultures, and for various buffer formulations are provided at the bottom of the page in the protocols tab.
Related Sections: Protein Electrophoresis Reagents Selection and Preparation
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.