Two-Dimensional Separation System by Coupling Capillary Isoelectric Focusing to Pressurized Capillary Electrochro-matography for Peptides Separation
A novel 2D platform combining capillary isoelectric focusing(cIEF)with pressurized capillary electrochromatography(pCEC)has been developed for off-line separation of peptides.The HPC coated capillary of cIEF was applied in the separation of standard proteins,and the average efficiency was around 310000.A homemade partition cell was induced in the end of cIEF capillary to make the collection of the eluting section more convenient.Compared to the capillary reversed phase liquid chromatography(CRPLC),the 6 kV voltage applied in the pCEC capillary enhanced the resolution of peptides,along with a faster velocity.The separation effectiveness of this cIEF/pCEC system was demonstrated using BSA tryptic digests.Theoretically,peak capacity of more than 30000 could be achieved.Moreover,this system was used for the separation of human red blood cell lysate digests.More than 200 peaks were detected in the 7 fractions from cIEF.The power of this system could be further enhanced by increasing the number of cIEF fractions.