The Quantification of Glycosaminoglycans: A Comparison of HPLC, Carbazole, and Alcian Blue Methods/title>

摘要

Glycosaminoglycans (GAGs) are linear polysaccharides that are found in the extracellular matrix and biologicalfluids of animals where they interact with hundreds of proteins and perform a variety of critical roles. There are fiveclasses of animal GAGs: heparan sulfate (HS), chondroitin sulfate (CS), dermatan sulfate (DS), keratan sulfate (KS), andhyaluronan (HA). Many biological functions can be monitored directly by their impact on GAG quantity. Thus, simple,sensitive, and robust GAG quantification methods are needed for the development of biomarkers. We have systematicallycompared three available GAG quantification assays including an HPLC-based assay, a simplified Alcian Blue assay, anda miniaturized carbazole assay. The carbazole and Alcian Blue assays were reproducible and simple to perform in generallab settings, but had important limitations: The carbazole assay could not detect KS and it overestimated GAGs that werecontaminated with salts or dissolved in PBS. The Alcian Blue assay detected only those GAGs that were sulfated. In contrast,while the HPLC method was time-consuming, it was a robust and sensitive assay that not only detected all GAGsbut also quantified glucosamine-GAGs and galactosamine-GAGs simultaneously. The HPLC assay was not affected bysalt or level of GAG sulfation and it yielded reproducible values for all types of GAGs tested. These results suggest thatan automated HPLC assay would be generally useful for the routine measurement of a panel of GAG-based biomarkerswhile the carbazole assay and the Alcian Blue assays could prove valuable for more specific purposes.