Basis of differentiation | Stacking gel | Separating gel |
Definition | Is a low concentrated polyacrylamide gel used in SDS-PAGE technique to concentrate molecules into a tight band, readying them for separation in the separating gel | Is a highly concentrated polyacrylamide gel used in SDS-PAGE technique to separate molecules based on size, revealing the composition of the sample |
% of polyacrylamide in the gel | Has a lower percentage of polyacrylamide - about 4%, leading to larger pore sizes | Has a higher percentage of polyacrylamide – around 10% resulting in smaller pore sizes and finer matrix |
Pore Size | Has larger pore size that enables rapid movement of proteins and helps proteins concentrate at the interface with the separating gel | Has smaller pore size that restricts the movement of proteins, promoting separation based on molecular weight |
pH | Has a lower pH of 6.8 | Has a higher pH of 8.8 |
Placement | Is placed on top of the separating gel | Is placed at the bottom of the gel electrophoresis container |
Resolution | Gives lower resolution | Gives better |
Function | To line up and stack proteins into a tight band, so they can enter the resolving gel at the same time | To separate proteins based on molecular weight |