AI Article Synopsis
- The chapter details a laboratory methodology for analyzing the N-glycome in cerebrospinal fluid (CSF) using various techniques.
- N-glycans are extracted from glycoproteins through digestion with PNGase F and purified using specialized solid phase extraction cartridges.
- The resulting glycans are permethylated for enhanced analysis via mass spectrometry, helping to link specific N-glycome profiles to different stages of Alzheimer's disease.
Article Abstract
In this chapter, we will present the methodology currently applied in our laboratory for the structural elucidation of the cerebrospinal fluid (CSF) N-glycome. N-glycans are released from denatured carboxymethylated glycoproteins by digestion with peptide-N-glycosidase F (PNGase F) and purified using both C18 Sep-Pak and porous graphitized carbon (PGC) HyperSep™ Hypercarb™ solid phase extraction (SPE) cartridges. The glycan pool is subsequently permethylated to increase mass spectrometry sensitivity. Molecular assignments are performed through matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI TOF MS) analysis considering either the protein N-linked glycosylation pathway or MALDI TOF MS/MS data. Each stage has been optimized to obtain high-quality mass spectra in reflector mode with an optimal signal-to-noise ratio up to m/z 4800. This method has been successfully adopted to associate specific N-glycome profiles to the early and the advanced phases of Alzheimer's disease (AD).
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| http://dx.doi.org/10.1007/978-1-0716-3774-6_4 | DOI Listing |
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