ஜர்னல் ஆஃப் அனலிட்டிகல் & பயோஅனாலிட்டிகல் டெக்னிக்ஸ்

சுருக்கம்

Assay for Glycosaminoglycans by Tandem Mass Spectrometry and its Applications

Shunji Tomatsu, Tsutomu Shimada, Robert W Mason, Joan Kelly, William A LaMarr, Eriko Yasuda, Yuniko Shibata, Hideyuki Futatsumori, Adriana M Montaño, Seiji Yamaguchi, Yasuyuki Suzuki and Tadao Orii

Glycosaminoglycans (GAGs) are distributed in the whole body and play a variety of important physiological roles associated with inflammation, growth, coagulation, fibrinolysis, lipolysis, and cell-matrix biology. Accumulation of undegraded GAGs in lysosomes gives rise to a distinct clinical syndrome, mucopolysaccharidoses. Measurement of each specific GAG in a variety of specimens is urgently required to understand GAG interaction with other molecules, physiological status of patients, and prognosis and pathogenesis of the disease.
We established a highly sensitive and accurate tandem mass spectrometry (LC-MS/MS) method for measurements of disaccharides derived from four specific GAGs [dermatan sulfate (DS), heparan sulfate (HS), keratan sulfate (KS), and chondroitin sulfate (CS)]. Disaccharides were produced by specific enzyme digestion of each GAG, and quantified by negative ion mode of multiple reaction monitoring. Subclasses of HS and GAGs with identical molecular weights can be separated using a Hypercarbcolumn (2.0 mm×50 mm, 5 μm) with an aectonitrile gradient in ammonium acetate (pH 11.0).
We also developed a GAG assay by RapidFire with tandem mass spectrometry (RF-MS/MS). The RF system
consists of an integrated solid phase extraction robot that binds and de-salts samples from assay plates and directly injects them into a MS/MS detector, reducing sample processing time to ten seconds. RF-MS/MS consequently yields much faster throughput than conventional LC-MS/MS-based methods.
However, the RF system does not have a chromatographic step, and therefore, cannot distinguish GAGs that
have identical molecular weights. Both methods can be applied to analysis of dried blood spots, blood, and urine specimens.
In this article, we compare the assay methods for GAGs and describe their potential applications.