A double-vented tetraphasic continuous column approach to MuDPIT analysis on long capillary columns demonstrates superior proteomic coverage.
A double-vented serial tetraphasic capillary column approach is applied to proteomic MuDPIT-type analysis using extended length capillary reverse-phase columns. The heart of the tetraphasic device consists of a triphasic MuDPIT trap located upstream of a venting tee. The trap is followed by a 60 cm high-resolution capillary column. A conventional high-flow HPLC is used to develop gradients at standard flow rates and pressures. The double-vented triphasic MuDPIT trapping device relieves the capillary separation column from the salt burden during the on-line cation-exchange portion of the analysis. Two configurations are presented, a double-vented continuous column model and a discontinuous model in which the triphasic MuDPIT trap is installed on a six-port valve; both configurations were tested with 60 and 10 cm capillary columns. All four systems were challenged with a trypsin digest of undepleted human serum, and a matrix of proteomic results for the different models and column lengths are compared.
Guzzetta
Andrew W
AW
Vincent Coates Foundation Mass Spectrometry Laboratory, Stanford University, Stanford, California 94305, USA. guzzetta@stanford.edu
Chien
Allis S
AS
D001798:Blood Proteins / Q000737:chemistry; D002412:Cations; D002845:Chromatography; D002851:Chromatography, High Pressure Liquid; D002852:Chromatography, Ion Exchange; D019075:Electrophoresis, Capillary; D006801:Humans; D013058:Mass Spectrometry; D036103:Nanotechnology; D010455:Peptides / Q000737:chemistry; D020543:Proteome; D040901:Proteomics / Q000295:instrumentation* / Q000379:methods*; D013997:Time Factors; D014357:Trypsin / Q000494:pharmacology
D001798:Blood Proteins; D002412:Cations; D010455:Peptides; D020543:Proteome; D014357:Trypsin