Lectin magnetic bead array for biomarker discovery.

Alterations in protein glycosylation play an important role in patho-physiology, and much effort has been devoted to detecting glycoprotein biomarkers. In this manuscript, we describe the development of a novel method for monitoring alterations in protein glycosylation. Lectins are used as individual affinity reagents and coupled to magnetic beads (Dynabeads) in a microplate array format for isolation of glycosylated proteins. Isolated glycoproteins are digested with trypsin in-solution followed by LC-MS/MS, allowing a liquid handler-assisted high throughput workflow. We demonstrate the specific and reproducible affinity-isolation of glycoproteins using the lectin Dynabead array technology. When used with serum, we achieved one-step purification of glycoproteins with minimal coisolation of abundant serum proteins including albumin. We further optimized the proteomics workflow to allow transfer to a liquid handler for automation. In summary, we report the development of a high throughput platform to detect alterations in protein glycosylation which will be useful in glycoproteomics studies, particularly clinical proteomics studies where large sample sizes are required to achieve statistical power.