1. Abstract This paper is focused on the study of different functionalised nanostructurated silicon – porous silicon – layers with the aim to find the suitable surfaces for protein immobilization. The remarkable advantages of this material include its huge surface to volume ratio, its high chemical reactivity at room temperature and potential compatibility with silicon integration technologies. The porous silicon is formed through an electro-chemical wet etching process, which can be tailored to control the pore depth, as well as the pore size and spacing. Prior to carrying out the electro-chemical etch; the bottom-surface of a boron doped p-type Si wafer is coated with a layer of metal (Al) for uniform voltage distribution across the wafer. In order to obtain different pore morphologies as well as different densities of active sites, besides tuning of process parameters, Si wafers with two crystallographic orientations has been used. Therefore, the starting material was Si with (111), and respectively (100) crystal orientation and resistivities of 10–20 Ωcm. The current density and concentration of electrolyte – hydrofluoric acid and ethanol based solution – were modified in order to obtain different pores size. The removing of any organic and inorganic impurities the as-prepared porous silicon (PS) has been achieved by boiling in HCl: HNO3 for 10 min. All PS test slides were printed with BSA dilutions labelled with Cy3 at 70% humidity and 22 o C in 30x48 probes/slide density. The slides were analyzed by laser excitation and the signal intensity for each type of surface has been recorded. The signal intensity is proportional to the quantity of protein (BSA), but the nanostructuration of surface decrease signal accuracy because the background is to higher. SEM, FTIR UV-VIS parallel investigations were performing for completed investigation of immobilization reaction. Porous Silicon Layer For Protein Immobilization M. Simion, M. Miu, I. Kleps, A. Bragaru, T. Ignat National Institute for Research and Development in Microtechnologies (IMT), Bucharest, 077190, Romania L. Ruta, I. Baciu Bucharest University, 00133 Bucharest, Romania C. Mihailescu, D. Stan DDS Diagnostic Bucharest, Romania e-mail: monica.simion@imt.ro; moni304ro@yahoo.commonica.simion@imt.romoni304ro@yahoo.com Conclusions The increased textured surface area allows the immobilization of more molecular species in the surface layer of a substrate as compared to a planar surface and thus obtaining an increased chemical activity. low surface background; intensity high immobilization efficiency; low surface wetting; lower limit of detection; Most important advantage of this substrate was the « coffee ring effect absence; Fluorescence intensity as a function of the protein target concentration on PS substrate, after printing, 24 h incubation and after 30 min washing SEM images of the PS samplesSurface Cross section Surface Cross section Array fluorescent image UV- VIS spectra of washing water FTIR spectrum of PS and PS with BSA imobilizeated