Mucins of the corneal and conjunctival epithelia are necessary for the protection of the ocular surface against desiccation, pathogen access, and injury. Detection and quantification of mucins is important for the understanding of ocular surface diseases that cause impaired vision and, in advanced stages, blindness. Advances in the field of molecular biology have made it possible to study membrane mucins and their associated O-glycans in established cell culture models of human ocular surface epithelia. This chapter discusses procedures to detect and quantify mucin RNA and protein in biological samples, as well as methods to experimentally manipulate the epithelia in culture by shRNA, to understand the function of specific mucins. Example protocols are provided to evaluate the role of ocular surface mucins in mucosal barrier function and bacteria-host interactions.