Charged surface reversed phase chromatographic materials method for analysis of glycans modified with amphipathic, strongly basic moieties

A method for selectively isolating glycans from a sample based on the net charge of each glycan, the method comprising:

• a) reacting a sample comprising glycans with a labeling reagent to produce labeled glycans;
• b) loading the labeled glycans onto a chromatographic separation device comprising a high purity chromatographic material comprising a chromatographic surface wherein the chromatographic surface is charged and comprises a hydrophobic surface group and one or more ionizable modifiers, such that the labeled glycans are selectively adsorbed onto the high purity chromatographic material;
• c) selectively eluting the adsorbed labeled glycans from the high purity chromatographic material, the selective elution including:
• exposing the labeled adsorbed glycans on the high purity chromatographic material to an aqueous mobile phase, the aqueous mobile phase comprising a concentration of a buffer and a concentration of an organic solvent, increasing the concentration each of the buffer and the organic solvent, thereby varying ionic strength of the aqueous mobile phase, and
• selectively separating the labeled glycans from the high purity chromatographic material based on the net charge of each glycan and the ionic strength of the aqueous mobile phase;
• wherein the organic solvent comprises water and acetonitrile and the buffer is selected from the group consisting of a first buffer comprising ammonium formate and formic acid, a second buffer comprising ammonium formate and acetic acid, and a third buffer comprising ammonium acetate and acetic acid.