Capillary electrophoresis is a powerful methodology for quantification and structural characterization of highly anionic polysaccharides. Separation of saccharides under conditions of electrophoretic flow, typically achieved under low pH (Ampofo et al., Anal Biochem 199: 249-255, 1991; Rhomberg et al., Proc Natl Acad Sci U S A 95: 4176-4181, 1998) is charge-based. Resolution of components is often superior to flow-based techniques, such as liquid chromatography. During the heparin contamination crisis, capillary electrophoresis was one of the key methodologies used to identify whether or not heparin lots were contaminated (Guerrini et al., Nat Biotechnol 26: 669-675, 2008; Ye et al., J Pharm Biomed Anal 85: 99-107, 2013; Volpi et al., Electrophoresis 33: 1531-1537, 2012).Here we describe a method for the isolation of sulfated heparin/heparan sulfate saccharides from urine, their digestion by deployment of heparinase enzymes (Ernst et al., Crit Rev Biochem Mol Biol 30: 387-444, 1995) resolution of species through use of orthogonal digestions, and analysis of the resulting disaccharides by capillary electrophoresis.
Keywords: Alcian Blue; Capillary electrophoresis; Heparan sulfate; Heparin; Heparinase; Pentosan polysulfate.
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