Journal
JOURNAL OF CHROMATOGRAPHY B-ANALYTICAL TECHNOLOGIES IN THE BIOMEDICAL AND LIFE SCIENCES
Volume 845, Issue 1, Pages 121-137Publisher
ELSEVIER SCIENCE BV
DOI: 10.1016/j.jchromb.2006.07.067
Keywords
lectin; multiple lectin; affinity chromatography; glycoproteins; mass spectrometry; human blood serum; high sensitivity
Funding
- NATIONAL CENTER FOR RESEARCH RESOURCES [P41RR018942] Funding Source: NIH RePORTER
- NATIONAL INSTITUTE OF GENERAL MEDICAL SCIENCES [R01GM024349] Funding Source: NIH RePORTER
- NCRR NIH HHS [RR018942] Funding Source: Medline
- NIGMS NIH HHS [GM24349] Funding Source: Medline
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We report here the use of high-performance lectin affinity enrichment of glycoproteins at microscale levels using a series of silica-bound lectins. The potential of this approach is being demonstrated for the glycoprotein enrichment from microliter volumes of human blood serum. Individual injections of sample to the affinity microcolumns packed with four lectin materials with different glycan specificities (Con A, SNA-I, UEA-I, PHA-L), followed by off-line reversed-phase pre-fractionation and nano-LC/MS/MS, permitted identification of 108 proteins in the lectin-bound fractions spanning a concentration dynamic range of 7-10 orders of magnitude. In contrast, multi-lectin microcolumn affinity chromatography, an alternative enrichment approach allowed identification of only 67 proteins. An attractive feature of high-performance lectin affinity chromatography at microscale levels is the substantial reduction of sample losses that are commonly experienced with extensive sample preparation needed for larger sample volumes. (c) 2006 Elsevier B.V. All rights reserved.
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