We performed experimental measurements and theoretical simulation based on an efficient half-space Green’s function method to investigate the diffraction patterns of light scattered from the biological structure on 1D reflection grating made of metal and polymer. The 1D grating provides higher-order reflected light, which can boost the image signal for off-specular reflection. This can facilitate the micro-ellipsometry imaging experiment when an incident angle of light is at a large angle, while the detection camera is placed at the upright position. The micro-ellipsometry images for s- and p-polarized reflectance and their phase difference (Rs, Rp, and Δ) was taken by a modified Optrel MULTISKOP system with rotating compensator configuration for various angles of incidence and wavelengths ranging from 450nm to 750nm. By using an 80X objective lens, the pixel size for our image is around 164nm. We can further increase the magnification and the numerical aperture by using a substrate collocated with a homemade acrylic resin lens, and the pixel size can be reduced to 50 nm. Based on the above, we study the optical properties of metallic/dielectric nanostructures and nearby biological systems including bacteria, and cancer cells via an imaging micro-ellipsometer combined with detailed theoretical modeling. By using specular and off-specular micro-ellipsometry imaging, we can achieve sufficient sensitivity to collect signals from a small area (around 10μm X 10μm and obtain a 3D image mapping of the morphology and dielectric properties of the biological system of interest.