4.2.2 Two-dimensional gel electrophoresis
Two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) separates proteins based on both mass and charge, allowing precise protein analysis. It can characterize around 5,000 distinct proteins depending on gel size. Proteins are dissociated based on their charge followed by their mass in first and second dimensions respectively. This technique is used for studying biochemical processes, post-translational modifications, and protein alterations [53]. The process involves five steps: sample preparation, first separation, linking, second separation, and protein identification. Isoelectric focusing usually precedes SDS electrophoresis. The traditional order is preferred for practical reasons, as SDS gels are easier to work with and compatible with downstream analysis methods like mass spectrometry [54].