Devices prepared by bottom-up strategies have been proposed for several biomedical applications. A product arising from this approach is often obtained from the self-assembly of a wide range of material classes. For instance, combining polymers showing distinct features or non-organic elements allow obtaining completely distinct properties useful not only for conventional healthcare but also for tissue engineering applications. Among various self-assembly techniques, layer-by-layer (LbL) has been introduced as an adsorption technique of elements with opposite charges into thin films. Lbl offers ease of preparation, versatility, fine control over the materials structure and robustness under physiological conditions. Due to its self-assembled nature and use of small masses of material (at the order of a few nanograms per layer), coatings can be produced by using aqueous solutions under mild conditions of temperature, pH and pressure, and minimal energetic requirements and material waste. In this chapter, we review the multitude of mechanisms present in the LbL technique for constructing nanostructured devices with interest for applications such as tunable cell adhesion and metabolism, drug delivery and biomineralized and smart systems.