Skip to content

How SDS-PAGE works

In the world of molecular biology and biochemistry, one technique reigns supreme when it comes to separating proteins by size: SDS-PAGE, or Sodium Dodecyl Sulfate Polyacrylamide Gel Electrophoresis. For years this method has provided a reliable means to analyze complex protein mixtures and determine their molecular weights. At its core, SDS-PAGE relies on a simple yet powerful principle – denaturation and uniform charge distribution. When proteins are mixed with SDS, a powerful detergent, they undergo denaturation, unraveling their complex three-dimensional structures and coating them with a negative charge. This uniform charge distribution ensures that proteins migrate through the gel primarily based on their size rather than their native charge, making SDS-PAGE a versatile tool for protein separation.

To further understand the mechanism behind SDS-PAGE, it’s crucial to delve into two key concepts: SDS binding and Stokes radius. SDS, as mentioned earlier, plays a pivotal role in this technique. By binding to the proteins and unfolding them into linear chains, SDS strips away their unique native structures and imparts a consistent negative charge along the length of the polypeptide chain. This negatively charged SDS-protein complex ensures that the rate of migration during electrophoresis is proportional to the protein’s size. The Stokes radius, on the other hand, represents the hydrodynamic size of a protein – essentially, how it behaves in a fluid environment. In SDS-PAGE, smaller proteins navigate through the acrylamide gel matrix with less hindrance, moving faster, while larger ones encounter more resistance and migrate more slowly. By exploiting these fundamental principles, scientists can separate proteins in a gel matrix, paving the way for a deeper understanding of complex biological systems and aiding in disease diagnosis, drug development, and countless other scientific endeavors.