Micro and nanofabrication technologies have been used extensively in many biomedical and biological applications. Integration of MEMS technology and biology (BioMEMS) enables precise control of the cellular microenvironments and offers high throughput systems. The focus of this research was to develop three dimensional (3-D) isotropic microstructures for comprehensive analysis on cell-substrate interactions. The aim was to investigate whether the normal and cancerous cells differentially respond to their underlying substrate and whether the differential response of the cells leads to a novel label-free technique to distinguish between normal and cancerous cells. Three different generations of 3-D isotropic microstructures comprised of curved surfaces were developed using a single-mask, single-etch step process. Our experimental model included HS68 normal human fibroblasts, MCF10A normal human breast epithelial cells and MDA-MB-231 metastatic human breast cancer cells. Primary findings on the first generation of silicon substrates demonstrated a distinct adhesion and growth behavior in HS68 and MDA-MB-231 cells. MDA-MB-231 cells deformed while the fibroblasts stretched and elongated their cytoskeleton on the curved surfaces. Unlike fibroblasts, MDA-MB-231 cells mainly trapped and localized inside the deep microchambers. Detailed investigations on cytoskeletal organization, adhesion pattern and morphology of the cells on the second generation of the silicon substrates demonstrated that cytoskeletal prestress and microtubules organization in HS68 cells, cell-cell junction and cell-substrate adhesion strength in MCF10A cells, and deformability of MDA-MB-231 cells (obtained by using AFM technique) affect their behavior inside the etched cavities. Treatment of MDA-MB-231 cells with experimental breast cancer drug, SAHA, on the second generation of substrates, significantly altered the cells morphology, cytoarchitecture and adhesion pattern inside the 3-D microstructures. Third generation of silicon substrates was developed for comprehensive analysis on behavior of MDA-MB-231 and MCF10A cells in a co-culture system in response to SAHA drug. Formation of colonies of both cell types was evident inside the cavities within a few hours after seeding the cells on the chips. SAHA selectively altered the morphology and cytoarchitecture in MDA-MB-231 cells. Most importantly, the majority of MDA-MB-231 cells stretched inside the etched cavities, while the adhesion pattern of MCF10A cells remained unaltered. In the last part of this dissertation, using AFM analysis, we showed that the growth medium composition has a pronounced effect on cell elasticity. Our findings demonstrated that the proposed isotropic silicon microstructures have potential applications in development of biosensor platforms for cell segregation as well as conducting fundamental biological studies. / Ph. D.