The increasing prevalence of diabetes and the severity of long-term complications have emphasized the need for continuous glucose monitoring. Optically-based methods are advantageous as they have potential for noninvasive or minimally invasive detection. Fluorescence-based affinity assays, in particular, can be fast, reagentless, and highly specific. Poly(ethylene glycol) (PEG) microspheres have been used to encapsulate such fluorescently labeled molecules in a hydrogel matrix for implantation into the body. The matrix is designed to retain the sensing molecules while simultaneously allowing sufficient analyte diffusion. Sensing assays which depend upon a spatial displacement of molecules, however, experience limited motility and diminished sensor response in a dense matrix. In order to overcome this, a process of hydrogel microporation has been developed to create cavities within the PEG that contain the assay components in solution, providing improved motility for large sensing elements, while limiting leaching and increasing sensor lifetime. For an implanted sensor to be successful in vivo, it should exhibit long-term stability and functionality. Even biocompatible materials that have no toxic effect on surrounding tissues elicit a host response. Over time, a fibrous capsule forms around the implant, slowing diffusion of the target analyte to the sensor and limiting optical signal propagation. To prevent this biofouling, a thermoresponsive nanocomposite hydrogel based on poly(N-isopropylacrylamide) was developed to create a self-cleaning sensor membrane. These hydrogels exist in a swollen state at temperatures below the volume phase transition temperature (VPTT) and become increasingly hydrophobic as the temperature is raised. Upon thermal cycling around the VPTT, these hydrogels exhibit significant cell release in vitro. However, the VPTT of the original formula was around 33-34 degrees C, resulting in a gel that is in a collapsed state, ultimately limiting glucose diffusion at body temperature. The hydrogel was modified by introducing a hydrophilic comonomer, N-vinylpyrrolidone (NVP), to raise the VPTT above body temperature. The new formulation was optimized with regard to diffusion, mechanical strength, and cell releasing capabilities under physiological conditions. Overall, this system is a promising method to translate a glucose-sensitive assay from the cuvette to the clinic for minimally invasive continuous glucose sensing.