This section details how to select and prepare the reagents (protein standards, gels, and buffers) required for various SDS-PAGE applications. The types of gels and buffers selected should suit the size of the protein under investigation, the desired analysis technique, and the overall goals of the experiment.
No particular gel type or buffer is useful for all proteins, and choosing the buffer systems and gel types that offer the highest resolution in the size range of interest may require some experimentation. In selecting reagents for SDS-PAGE, consider the following:
- Protein standards — select protein standards that provide maximum resolution in the size range of interest and that offer compatibility and utility for downstream applications such as western blotting
- Gel percentage — choose the percentage that offers resolution in the range of interest
- Precast vs. handcast gels — precast gels offer greater convenience and superior quality control and reproducibility than handcast gels; handcast gels offer the possibility of customized percentages and gradients
- Gel format — select mini- or midi-format gels when throughput is important or sample size is limited; select large-format gels for higher resolution. Select a comb type and gel thickness to accommodate the sample number and volume you are working with
- Buffer system — choose the system that offers the best resolution and compatibility with the protein and application of interest