Flow field-flow fractionation (FlFFF) has been successfully utilized for the size fractionation of macromolecules, nanoparticles, large biomolecules like proteins, DNA, subcellular organelles, biological vesicles, and cells. The size fractionation capability of FlFFF for biomaterials has led its utility toward proteomic and lipidomic applications in combination with liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS).
This presentation will show that FlFFF can be utilized for the separation of glycoproteins and membrane proteins from cancer patient samples, and the collected protein fractions can be analyzed by nLC-ESI-MS/MS for proteomics. Applications include proteomic and lipidomic analysis of lipoproteins in human blood plasma using off-line and on-line combination with MS. It demonstrates that high density lipoprotein (HDL) and low density lipoprotein (LDL) from patients plasma having coronary artery disease (CAD) can be isolated in their intact forms on a semi-preparative scale using multiplexed hollow fiber FlFFF and that collected fractions can be used for the lipidomic analysis of various lipids with structural identification using nLC-ESI-MS/MS. It will also show a recent development of on-line chip-type asymmetrical FlFFF with MS (AF4-ESI-MS/MS) for the top-down lipidomic analysis of human lipoproteins, demostrating that various phospholipids, regioisomers of triacylglycerols, and cholesteryl esters can be on-line analyzed which can be potentially utilized for the high speed screening of target lipid species in lipoproteins depending on disease status.