This dissertation is devoted to the development of novel bio-nano chip microfluidic platforms suitable for agarose bead-based sensors. Previously, our lab had developed a novel immunoassay sensor based on three-dimensional agarose beads whose surface is marked by micro-size pores. Porous agarose beads have a greater capacity of immobilizing antibodies as compared to other surface based sensors, and thus are expected to yield better assay performance. The first generation of bio-nano-chip platform had been designed to contain an array of three-dimensional wells to host the agarose beads. In the bio-nano-chip platform, a silicon-based bead holder had been designed to generate strong convectional flows around beads and greatly enhance the detection sensitivity. As the silicon microchip was fabricated based on the traditional MEMS techniques, however, the cost of the device remains relatively high. Moreover, the accuracy of the assays and the assay time also need to be improved to meet the standard of point-of-care diagnostics. In efforts to overcome these issues, this dissertation reports works of replacing the material of key components with cheaper materials in the bio-nano-chip platform. Further, this dissertation presents a new mechanism of recirculating assays in a membrane-based chip, and also a new bead design to further enhance the assay performance. In summary, the bio-nano-chip platform had been improved to be more efficient and cost-effective than previous designs, allowing wider applications in resource limited areas such as in developing countries.