An important tool for the performance analysis and standardization of medical imaging technologies is the phantom, which offers specifically defined properties that mimic the structural and optical characteristics of a tissue of interest. The development of phantoms for high-resolution (i.e., micro-scale) three-dimensional (3D) imaging modalities can be challenging, however, as few manufacturing techniques can capture the architectural complexity of biological tissues at such scales. Direct Laser Writing (DLW) is an evolving additive manufacturing technique with nano-scale precision that can fabricate micro and nanostructures with unparalleled geometric complexity. This dissertation outlines the unique microfluidic-based DLW strategies that have been developed for novel micro-scale phantom production. First, I will outline the development and characterization of an in situ DLW strategy used to adhere printed components to the surfaces of a microchannel. I will then explain how we have leveraged this strategy for a proof-of-concept retinal cone outer segment phantom that is laden with light-scattering Titanium (IV) Dioxide nanoparticles. This phantom has valuable implications for the performance analysis of the emerging ophthalmological modality, adaptive optics-optical coherence tomography (AO-OCT). Next, I will describe the development and characterization of a microfluidic-based multi-material DLW strategy to fabricate single components from multiple materials with minimal registration error between the materials. Ultimately, we intend to use this method to develop multi-material platforms and phantoms, including high-aspect-ratio multi-material retinal cone phantoms for AO-OCT. Finally, to demonstrate the applicability of this method for applications beyond AO-OCT, I present a preliminary phantom production strategy for the light microscopy-based modality, whole slide imaging (WSI). Specifically, we assess the DLW and light microscopy performance of dyed photoresists and offer a preliminary multi-material demonstration, which are pivotal first steps toward the creation of a first-generation multi-material WSI phantom. This work provides valuable insights and strategies that leverage microfluidic-based DLW techniques to fabricate novel micro-scale phantoms. It is anticipated that these strategies will have a lasting impact, not only on the production of phantoms for medical imaging modalities, but also for the fabrication of advanced microfluidic and multi-material microstructures for fields such as meta-materials, micro-optics, lab-on-a-chip, and organ-on-a-chip.