SDSPAGEvarianten

SDS-PAGE is a widely used method for separating proteins by molecular weight under denaturing conditions. Variants of SDS-PAGE refer to changes in gel composition, buffers, or sample preparation designed to improve resolution for certain protein classes or experimental needs. The core principle—SDS denatures proteins and imparts a roughly uniform negative charge, allowing separation by size in a polyacrylamide gel—remains the same.

Gel composition and format: common variants use fixed percentages (for example 8–12% acrylamide) or gradient gels (4–20%). Gel format can be the classic vertical Laemmli system or precast mini-gels. Stacking gels concentrate proteins before separation, improving lane consistency and resolution.

Buffer systems: the traditional Laemmli system uses Tris-Glycine-SDS buffers. Alternatives include Tris-Tricine-SDS, which improves resolution for small proteins, and MOPS- or Bis-Tris-based SDS buffers for different pH and running characteristics. These systems can affect separation, running time, and compatibility with staining methods.

Reducing versus non-reducing: many variants run samples under reducing conditions with DTT or beta-mercaptoethanol to break disulfide bonds, altering mobility when multimeric forms exist. Non-reducing variants preserve disulfides. Sample preparation may also involve heating with loading dye; some protocols omit heat for particular proteins or complexes.

Two-dimensional PAGE and other adaptations: A major variant is two-dimensional PAGE, in which an initial isoelectric focusing step is followed by SDS-PAGE. This variant dramatically increases resolving power and is used in proteomics. Other adaptations include SDS-PAGE with urea to enhance denaturation of hydrophobic proteins.

Choosing a variant depends on protein properties, desired resolution, and available equipment.