Developing microdevices on flexible material attracts scientific community to explore applications in different aspects of health and point of care diagnostics. Flexible substrates offer unique characteristics such as flexibility, stretchability, portability, low-cost, and simple fabrication. Fabrication of cost-effective paper-based analytical devices by wax printing has recently become popular using cellulose filter papers. Paper-based devices need higher temperature to form hydrophobic barrier across paper substrate, rely on large working channels (≥ 500 μm) for liquid handling, and exhibit lower efficiency (~50%) of sample mobility. Such limitations confine applications of wax-based fabrication. In this dissertation, we report printability, fidelity, and applications of wax micropatterns on polyethylene terephthalate-based substrate (PET), which is a a non-cellulosic, non-fibrous, and non-porous material. Resolution, sustainability against heat and biocompatibility was tested on wax micro-features. The patterned devices were explored for variety of applications.First, wax microwells on PET showed mouse embryonic stem cell (mESC) self-renewal or direct differentiation. Second, microfluidic flow was demonstrated on wax printed microchannels on PET which was used to develop distance-based assay. Third, fluidic properties of trinucleotide repeat sequences were investigated on wax microchannels. Fourth, multilayer wax-on-plastic device was fabricated using wax printing with hand painting of conductive materials for electrochemical immunosensing.