Zirconium(IV)-IMAC for phosphopeptide enrichment in phosphoproteomics

IMAC and MOAC magnetic microsphere materials were tested with ug amounts of protein extracts from HepG2/C3A cells. We compared Zirconium (IV) IMAC (Zr-IMAC) magnetic microspheres to commonly used Titanium (IV) IMAC (Ti-IMAC) and TiO2 magnetic microspheres for phosphopeptide enrichment from simple and complex protein samples prior phosphopeptide sequencing and characterization by mass spectrometry (LC-MS/MS). We optimized sample-loading conditions to increase phosphopeptide recovery for Zr-IMAC, Ti-IMAC and TiO2 based workflows. Furthermore, optimized Zr-IMAC and Ti-IMAC magnetic microsphere format were compared to HPLC-based Fe(III)-IMAC for same sample amount (200 ug).