Combining lectin affinity chromatography and immunodepletion – A novel method for the enrichment of disease-specific glycoproteins in human plasma

Drastic enrichment of potential disease-specific glycoprotein markers in human plasma can be achieved by the combination of affinity- and immuno-depletion. In the affinity-fractionation step all glycoproteins carrying a certain glycostructure are isolated by lectin affinity chromatography, thus depleting other components. Against the respective glycoprotein fraction isolated from the plasma of healthy individuals antibodies are raised in llamas. The llama heavy chain antibodies (which are particularly stable) directed at the isolated plasma glycoprotein fraction are immobilized and the immunoaffinity column thus obtained is used to deplete the respective glycoprotein fraction of patient plasma samples. Depletion of proteins normally found in human plasma by 99.8–99.9% can be achieved, resulting in a 800–1000-fold enrichment of potential disease-specific proteins in the flow-through of the immunoaffinity column.