EPSCoR Research Fellows: NSF: Developing LC-MS/MS and Immunofluorescence Workflows for Quantitative and Spatial Analysis of Glycosaminoglycans in the Sclera

This Research Infrastructure Improvement (RII) EPSCoR Research Fellows project provides a fellowship to an Assistant Professor and training for a graduate student at the University of Mississippi. This work is conducted in collaboration with Umesh Desai at Virginia Commonwealth University. Through the fellowship, the Principal Investigator will learn specialized methods for measuring sugar-like molecules called glycosaminoglycans in the sclera, the eye's tough outer wall, and map where they occur. The sclera holds the eye's shape and resists internal pressure. Its strength comes from collagen fibers, but sugar-like molecules control how those fibers are spaced and how the tissue bears load. Which types occur where is unknown. Drawing on biomedical engineering, analytical chemistry, and glycoscience, the project will combine chemical and imaging methods to find out, using pig eyes as a close model for the human eye. The findings will help explain why nearsightedness and glaucoma damage some eye regions more than others. The project will also build lasting measurement capability at the University of Mississippi, train students across disciplines, and strengthen statewide research infrastructure. This EPSCoR Research Fellows project will establish and validate two complementary workflows for class-resolved analysis of glycosaminoglycans (GAGs) across distinct scleral regions in a porcine model. It will deliver the first region-by-region account of which GAG classes occur where, which bulk assays cannot provide. Objective 1 will develop a liquid chromatography–tandem mass spectrometry (LC–MS/MS) workflow quantifying chondroitin, dermatan, heparan, and keratan sulfate and hyaluronan from enzymatically released disaccharides, validated for linearity, recovery, matrix effects, and reproducibility. Objective 2 will develop an immunofluorescence workflow mapping the same classes in situ using enzyme-validated antibodies and probes with blinded image analysis. Host-site training will reposition the PI's ocular biomechanics program toward glycoscience and will train a graduate student across analytical chemistry, imaging, and matrix biology. Both workflows will be implemented at the home institution as durable shared capability, integrated into an existing Research Experiences for Undergraduates (REU) program and biomechanics courses, and will sustain a new interdisciplinary partnership. This project is supported by the EPSCoR Research Infrastructure Improvement Program: EPSCoR Research Fellows (ERF). The ERF program supports early- and mid-career investigators in eligible jurisdictions to develop collaborations at the nation’s private, government or academic research institutions This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.