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Purpose: This study aims to develop biodegradable and biocompatible polymer-based nanofibers that continuously monitor pH within microenvironments of cultured cells in real-time. In the future, these fibers will provide a scaffold for tissue growth while simultaneously monitoring the extracellular environment., Methods: Sensors to monitor pH were created by directly electrospinning the sensor components within a polymeric matrix. Specifically, the entire fiber structure is composed of the optical equivalent of an electrode, a pH-sensitive fluorophore, an ionic additive, a plasticizer, and a polymer to impart mechanical stability. The resulting poly(ε-caprolactone) (PCL) and poly(lactic-co-glycolic acid) (PLGA) based sensors were characterized by morphology, dynamic range, reversibility and stability. Since PCL-based nanofibers delivered the most desirable analytical response, this matrix was used for cellular studies., Results: Electrospun nanofiber scaffolds (NFSs) were created directly out of optode material. The resulting NFS sensors respond to pH changes with a dynamic range centered at 7.8 ± 0.1 and 9.6 ± 0.2, for PCL and PLGA respectively. NFSs exhibited multiple cycles of reversibility with a lifetime of at least 15 days with preservation of response characteristics. By comparing the two NFSs, we found PCL-NFSs are more suitable for pH sensing due to their dynamic range and superior reversibility., Conclusion: The proposed sensing platform successfully exhibits a response to pH and compatibility with cultured cells. NSFs will be a useful tool for creating 3D cellular scaffolds that can monitor the cellular environment with applications in fields such as drug discovery and tissue engineering.