Glycoproteins are extremely important to a variety of biological processes, including protein folding and trafficking, cell-cell and cell-matrix interaction, cellular differentiation, fertilization, the immune response, and the initiation and metastasis of tumors. However, the analysis of glycoproteins is a big challenge because of the low abundance, the complexity of the glycan structures, the multiple substitutions (microheterogeneity) at glycosylation sites, and the structural diversity associated with the protein backbone itself.
There are a number of glycoprotein detection by staining procedures that can be used to detect highly glycosylated proteins on SDS gels, among which, periodic acid-Schiff (PAS) staining is the simplest and least expensive way to estimate whether a protein is glycosylated.
What Is PAS Staining?
The PAS staining method and its numerous variations are usually used to detect polysaccharides and carbohydrate macromolecules (such as glycoproteins, glycolipids, and proteoglycans) following sodium dodecyl sulfate (SDS) or non-denaturing polyacrylamide gel electrophoresis (PAGE) and protein transfer to nitrocellulose membranes. During the staining procedures, periodic acid is used to bind to the sugar groups of glycoproteins to overcome the problem that glycan sugar moieties are not reactive to staining or labeling molecules. The periodic acid oxidizes vicinal hydroxyls on sugars (especially sialic acid) to aldehydes or ketones, which reacts with the Schiff reagent to give a magenta color.