The paper dwells upon the elaboration of a simple and energy-efficient approach for the “bottom-up” fabrication of porous polymeric film, which possesses unique surface, structural and bulk properties, followed by its comprehensive study through various characterization techniques. The fabricated film is multi-layered methyl silsesquioxane and suitably modified with the help of several treatments making the film compatible for biosensing applications. Film characterization reports the self- organized topography of the film deposited over the substrate and confirms the successful bonding between the polymers. The influence of the concentration of polypropylene glycol, which itself acts as thermally labile dendrimer, is studied by observing its bond formation ensuring polymer mixing by using FTIR. Energy dispersive x-ray analysis has been performed for the confirmation of elements. In addition to this, the nanoporous film is also studied with the help of variable angle spectroscopic ellipsometry for the determination of film porosity and refractive index. In this analysis, Cauchy model and graded-index model were chosen for data analysis and the Bruggeman effective medium approximation method was used for evaluating the optical constant. The surface of the porous film has been treated with different silane surfaces, characterized with the help of FTIR, UV-VIS, and AFM and envisioned for protein immobilization. © 2021 The Authors