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Rapid separation and characterization of protein and peptide mixtures using 1.5 μm diameter non‐porous silica in packed capillary liquid chromatography/mass spectrometry
Author(s) -
MacNair John E.,
Opiteck Gregory J.,
Jorgenson James W.,
Moseley M. Arthur
Publication year - 1997
Publication title -
rapid communications in mass spectrometry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.528
H-Index - 136
eISSN - 1097-0231
pISSN - 0951-4198
DOI - 10.1002/(sici)1097-0231(199708)11:12<1279::aid-rcm18>3.0.co;2-s
Subject(s) - chemistry , chromatography , capillary action , mass spectrometry , characterization (materials science) , packed bed , porosity , peptide , analytical chemistry (journal) , organic chemistry , nanotechnology , biochemistry , materials science , composite material
Octadecyl‐modified 1.5 μm diameter non‐porous silica particles were packed in 150 μm i.d. (360 μm o.d.) capillaries with lengths of 20 cm which were used to separate proteins and peptides generated from enzymatic digests of proteins. Gradients were produced using an exponential dilution method at pressures of 520 Bar (7500 psi) and electrospray ionization mass spectrometry was used for detection. This system was similar to packed capillary perfusion chromatography with respect to chromatographic resolution and analysis time and had a limit of detection comparable to traditional packed capillaries which use 5 μm diameter porous particles. The analyses required as little as 250 femtomol of protein or 500 femtomol of peptide on‐column in approximately 30 min. This technique was then applied to verify the existence of an overexpressed protein in an E. coli cell lysate and to confirm the presence of four glycoforms of a peptide generated in the proteolytic digest of an antibody. © John Wiley & Sons, Ltd.