The ocular disease Glaucoma affects around 3 million people in the United States, and serves as the second leading cause of blindness. The primary structure effect is known as the Trabecular Meshwork (TM), similar to an adaptive filter as its permeability alters depending on the pressure inside the eye. Aqueous humor flows through the TM, nourishing the surrounding tissues with glucose and oxygen. In patients with Glaucoma, overexpression of certain genes causes the TM to slowly stiffens with a reduction in aqueous humor outflow. Current glaucoma medications work by lowering intraocular pressure (IOP), a risk factor for glaucoma, through either decreasing production of aqueous humor or increasing the permeability of the trabecular meshwork. However, these treatments only mitigate the symptoms of the disease, not addressing the root cause of gene overexpression. In previous studies, two specific glaucoma genes, SPARC and TGFB2, have been associated with stiffening the extracellular matrix of the trabecular meshwork, leading to an increase in intraocular pressure. In my research, RNAscope was performed to validate and quantify the exact location of gene expression between normal and glaucoma donor eyes. The experiment also pioneered the procedure of duplex RNAscope to efficiently analyze two different genes at the same time using the same tissue. In the laboratory, specific mRNA probes were used to bind to the RNA sequence inside of the cell. In order to better visualize the signal, amplifiers increased potential attachment sites for the colored staining solutions. There was significant overexpression of both genes in the cornea, retina, and optic nerve structures. However, the genes had low expression in the trabecular meshwork, with slightly increased expression in glaucomatous eyes. The duplex RNAscope was very successful and can be utilized to save precious human donor tissue. The process also aids in preventing any potential biological or structural variation between donor tissue. The research progresses the field towards greater ideal glaucoma therapy solutions, treating the physiological processes with long term improvements. Further research is needed to quantify the exact value of overexpression seen between the two categories, and to test for more glaucoma associated genes.