GlycoTechnology

The Glycotechnology Core Resource utilizes the following instrumentation for routine analysis:

1. A Dionex ICS 3000-HPLC equipped with a electrochemical and conductivity detector, dual gradient pump and AS 50 autosampler. It is primarily used for the composition analysis of monosaccharides, uronic acids, sialic acids after separation by high pH anion exchange chromatography with pulsed amperometric detection and for the analysis of cations using conductivity detection. It is also used for N+O profiling of released oligosaccharides. Collection of appropriate fractions for further analysis is also possible.

2. A Dionex DX-500 HPLC equipped with a GP-40 quaternary gradient pump, a GP-50 quaternary gradient microbore pump, a ThermoSeparations AS3500 autosampler, an LC20 chromatography enclosure, an AD20 variable wavelength UV detector and an ED40 pulsed amperometric/conductivity detector is utilized primarily for the compositional analysis of neutral and amino sugars; uronic acids; and sialic acids following separation by high pH anion exchange chromatography with pulsed amperometric detection. Fraction collection is possible.

3. A Dionex DX-600 BioLC equipped with a GP-50 quaternary gradient pump, a ThermoSeparations AS3500 and a Dionex variable wavelength fluorescence detector is primilarly utilized for the analysis of sialic acids by reverse phase chromatography following derivitization with DMB with fluorescence detection. This system is also used for the analysis of 2-AA or 2-AB tagged oligosaccharides by normal phase separation on an amine-bonded column with fluorescence detection. Fraction collection is possible.

4. A Dionex DX-600 BioLC equipped with a GP-50 quaternary gradient pump, AS-50 autosampler, LC30 chromatography enclosure, AD 25 variable wavelength UV detector an ED50 pulsed amperometric/conductivity detector and a Jasco 1520 fluorescence detector, a column heater, an Eldex dual channel post-column addition pump, and a heated post-column reactor is primarily used for the analysis of glycosaminoglycan-derivied disaccharides enzymatically generated by the action of lyases. Detection is based on absorbance at 232 nm and on post-column fluorescent derivatization using 2-cyanoacetamide.

A Hewlett-Packard HP5890 series II gas chromatograph with a HP5971A mass selective detector is used for monosaccharide analysis following hydrolysis and TMS derivatization and for linkage analysis of partially methylated alditol acetates.

A ThermoFinnigan LC-Q ion-trap mass spectrometer with associated HPLC comprised of a ThermoSeparations AS3500 autosampler, a P4000 quaternary gradient pump and UV6000LP diode array detector is used for electrospray ionization, ion trap mass spectrometric determinations of analyte masses.

The instrument is a hybrid quadrapole time-of-flight mass spetrometer, a configuration ideally suited for the analysis of all classes of glycoconjugates including free glycans, glycopeptides, and glycolipids, and for protein identification and peptide sequencing. Also, the Applied Biosystems QSTAR XL QqTOP software controls both the Agilent 1100 and the LC Packings Ultimate HPLC systems.

Instrumentation