Negatively charged glycans (sialic acids, sulphated or phosphorylated sugars) often play a critical role in the function of a glycoprotein.
Sialic acids increase the serum half-life of glycoproteins by protecting them from degradation by the asialoglycoprotein receptor; sulphated glycans are involved in cell adhesion; and Mannose-6-Phosphate is a key targeting signal for transport of glycoproteins to lysosomes and is present in therapeutic enzymes (enzyme replacement therapies) developed for treatment of lysosomal storage diseases.
The LudgerSep-C3 column is a weak anionic exchange (WAX) HPLC column that enables you analyse negatively charged sialylated, phosphorylated and sulphated glycans. This technique is also known as ‘charge profiling’.
An example of the information that can be provided is the relative amounts of sialylation (1, 2, 3 or 4 sialic acids) on your glycoprotein which is important to know when analysing a highly sialylated protein such as erythropoietin (EPO).
Although sialylated and sulphated glycans can be separated by anion exchange at a low pH of 4.4, the phosphorylated sugars would not be fully charged and there would be multiple species in solution.
In order to have one buffer which is suitable for separation of all anionic glycans (sialic acids, phosphorylated and sulphated sugars) Ludger recommends the use of pH9 ammonium formate buffer.
The LudgerSep C buffer is a pH 9 ammonium formate buffer concentrate which can easily be diluted with water and acetonitrile then used directly as a solvent for the LudgerSep C3 column.
