Traditional methodologies to study in vitro biological processes include simplified laboratory models where different parameters can be measured in a very controlled environment. The most used of these practices is cell plate-culturing in aqueous media. In this minimalistic model, essential components of the biological system might be ignored. One of them, disregarded for a long time, is the extracellular matrix (ECM). Extracellular matrix in eukaryotic cells is not only a frame for cells and biological components, but also an active partner of cellular metabolism and participates in several normal and pathological biological processes in a dynamic manner. ECM of eukaryotic cells has a very complex structure. Also, its mechanical properties (stiffness, viscoelasticity) depend on the organ it is associated with, and may vary from a very fluid (plasma) to a very solid (bones) structure. ECM structure and composition are very dynamic and experience temporal structural and topological changes, affecting all the existing interactions. When mimicking the ECM, three aspects are considered: the chemical environment and the physical and structural properties. In this review, we present two lines of research studying the role of the ECM in two biological implications: membrane fluidity heterogeneity and protein retention and aggregation. For these studies, we used biopolymeric matrices with very controlled features to evaluate the two events. We use traditional biochemical techniques and fluorescence microscopy to study the biological systems and traditional polymer techniques (rheology, SEM) to characterize the polymeric matrices.