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Updated: Jan 7, 2026
Sample Preparation for Mass-spectrometry-based Proteomics Analysis of Ocular Microvessels
Published on: February 22, 2019
Keratan sulfate revisited: UPLC-MS/MS-based quantitative profiling reveals structural heterogeneity and deficiency in
Xinping Wang1, Han Zhou1, Xi Liang1
1Key Laboratory of Marine Drugs of Ministry of Education, Shandong Key Laboratory of Glycoscience and Glycotherapeutics, School of Medicine and Pharmacy, Ocean University of China, Qingdao, 266003, China.
None:
Keratan sulfate (KS) is a structurally unique glycosaminoglycan involved in various physiological processes. Dysregulated KS levels and sulfation modifications are associated with the development of various disorders. However, the absence of commercial KS standards and specific hydrolases has hindered accurate quantification and functional studies. To overcome these limitations, we prepared two structurally distinct KS disaccharide standards (Gal-GlcNAc6S and Gal6S-GlcNAc6S) from corneal and cartilaginous tissues, and obtained recombinant KS hydrolase through heterologous expression. Given that the total abundance of various sulfated KS disaccharides reflects overall KS content, a robust UPLC-MS/MS method was further developed for simultaneous quantification of multiple sulfated KS disaccharides. The developed methodology demonstrated exceptional sensitivity, achieving KS quantification from minimal biological inputs (500-1,000 cells or 1 mg tissue). Quantitative analysis revealed significant variation in KS distribution among distinct human ocular cell populations, with stromal cells exhibiting the highest abundance, followed by endothelial and epithelial cells. Notably, total KS expression in corneal specimens from macular corneal dystrophy (MCD) patients was reduced by over 90 % relative to keratoconus (KC). This study establishes a comprehensive platform for KS quantification and offers novel insights into its structure-function relationships within ocular tissues.
