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Mucus is a biological hydrogel which lines the wet (non-keratinized) epithelia of the body. Mucus provides a gateway between the cells of the epithelium and the outside world, and is postulated to provide a selective filtering function which is critical to physiological functioning and has been implicated in diseases. Currently, much of the mechanisms and criteria of this selective filtering function is not well understood. In this thesis, we contribute novel microfluidic devices to characterize the selective permeability properties of the mucus barrier. Microfluidics provides the engineering ability to create channels with precise geometries, fluid flow capability, and allow chemical concentration gradients. Our devices mimic the physiological environment of the mucosa and enable improved measurements of the mucus layer selective permeability. The first microfluidic device mimics the acid barrier function of the stomach mucus layer. This device reproduces on-chip the secretion of mucus by the gastric mucosa into an acidic stomach lumen. We use this device to demonstrate that the secretion of mucins, the glycoprotein structural component o