Intact liposomes have been immobilized onto solid surfaces by a NeutrAvidin-biotin link. The construction of these layers has been followed up by X-ray photoelectron spectroscopy (XPS) and quartz crystal microbalance (QCM) measurements with energy dissipation monitoring. Also, the simultaneous release of two fluorescent probes from these liposome layers has been investigated with the aim to validate this method in multirelease delivery systems. XPS showed the successful immobilization of the different layers. XPS results also point out the importance of the deactivation method used to reveal the presence of the specific NeutrAvidin-biotin attachment. QCM measurements allowed the buildup of the different layers to be followed in real time and in situ and suggest that biotinylated liposomes stay intact upon surface attachment on NeutrAvidin-covered surfaces and had viscoelastic behavior. QCM experiments also demonstrated that surface-immobilized liposomes were able to resist irreversible adsorption from fetal bovine serum. Release kinetic profiles were studied by monitoring the release of two different fluorescent probes, namely, carboxyfluorescein and levofloxacin, from these liposome layers. These studies showed that it was possible to modulate to some extent the release rates of the two molecules by using different configurations of liposome layers.