A Spin-Tip Enrichment Strategy for Simultaneous Analysis of N-Glycopeptides and Phosphopeptides from Human Pancreatic Tissues

The dual-functional Titanium immobilized metal affinity chromatography strategy can enrich both N-glycopeptides and phosphopeptides in the same workflow, increasing biomolecular information from the same analysis. The main advantage of this enrichment is two separation mechanisms, which enable the simultaneous separation of N-glycopeptides and phosphopeptides in separate fractions for downstream mass spectrometry analysis. Using this enrichment method can improve detection of glycosylation and phosphorylation in complex biological samples like pancreatic tissues toward disease biomarker discovery such as in diabetes and cancer.

Because this method relies on spin tips and centrifugation, it is important to control the centrifugation speed, and make sure that samples are free of particulates to prevent any clogging. To begin, pre-chill the parts of the tissue pulverizer that will come in contact with the pancreatic tissues, namely the chamber, pulverizer, and recovery spoon, along with any spatulas, in a polystyrene container. Next, transfer tissue pieces into the pre-chilled sample holder, and add a liquid nitrogen to the tissue until they are frozen.