- Langmuir–Blodgett (LB) films of the conjugated polydiacetylene (PDA) exhibit spectroscopic behavior, which is dependent on the type of the supporting substrate. While on polished silicon surfaces the photoluminescence (PL) of PDA is quenched, it is preserved on top of 2D patterned macro-porous silicon (2D-MPS). 2D-MPS, prepared by electrochemical etching of photo-lithographically pre-patterned silicon, is a 2D array of ca. 10 μm deep pores with lateral 2–4 μm repeating unit cells in orthogonal or hexagonal arrangements. LB films of PDA on such surfaces form membranes with continuous domains of the size sufficient to cover laterally many cell units. Apparently, the PL from this film results exclusively from the portion of the PDA membrane which is suspended over pore openings, while portions of the film which are attached to the silicon on top of the pores walls does not exhibit PL at all. We have used these membranes in different configurations and exposed them to different chemical and biological agents and followed the PL intensity change. This report demonstrates the effectiveness of the combined system: LB films of PDA on top of 2D-MPS as sensing probe for a variety of chemicals including, Cd ions and TNT explosives. In addition, the use of films of PDA, in which glycol-lipid were embedded, for binding and recognition of lectin protein, mimicking the cell membrane interaction with its environment, is also demonstrated.