Abstract
Purpose :
To utilize a novel capillary electrophoresis nano-immunoassay system to evaluate and compare regional conjunctival protein expression of ocular surface mucin MUC1 in normal and dry eye subjects.
Methods :
Adults with and without signs and symptoms of dry eye disease were recruited. Impression cytology was performed by placing a semicircle of filter paper on the bulbar conjunctiva to collect epithelial cells and membrane associated mucin MUC1 protein. Cells were collected from the temporal, superior, inferior, and nasal bulbar conjunctiva of both eyes, and the lid wiper area of the upper eyelids. After protein extraction using RIPA buffer, two samples from both eyes of a single individual were pooled into one protein sample and expression was analyzed using Jess, a capillary electrophoresis nano-immunoassay system. The chemiluminescence intensity of each single antigen binding signal was calculated using the Compass Simple Western software 6.1 following normalization to the total protein amount. Statistical analyses were conducted using GraphPad Prime 9.
Results :
Seven dry eye and seven normal subjects were enrolled. Identifiable antibody binding signals for MUC1 were seen in all samples around 280 kDa as expected. In normal samples, MUC1 protein expression from the nasal bulbar conjunctiva was significantly greater than the superior (p=0.004), inferior (p=0.005), and eyelid margin (p=0.008). While not statistically significant, the MUC1 binding signal intensity was also higher in the temporal bulbar conjunctiva. In the dry eye samples, MUC1 expression was highest in the superior bulbar conjunctiva compared to the inferior (p=0.01) and nasal (p=0.001) regions. When comparing MUC1 expression between dry eye and normal subjects, MUC1 expression was lower in the nasal (p<0.0001) and temporal (p=0.01) regions of the dry eye samples.
Conclusions :
The Jess system provides numerous advantages over a traditional Western blot especially when analyzing high molecular weight proteins like ocular surface mucins with small sample aliquots. The increased expression of MUC1 in the temporal and nasal bulbar conjunctiva of healthy subjects suggests a functional need for increased lubrication in these areas not normally protected by the eyelids. In addition, the decreased MUC1 expression in dry eye samples, especially temporally and nasally, is consistent with results from previous studies in dry eye patients.
This abstract was presented at the 2022 ARVO Annual Meeting, held in Denver, CO, May 1-4, 2022, and virtually.