In the present work, a technique for establishing multiple black lipid membranes (BLMs) in arrays of micro structured ethylene tetrafluoroethylene (ETFE) films, and supported by a micro porous material was developed. Rectangular 8 × 8 arrays with apertures having diameters of 301 ± 5 µm were fabricated in ETFE Teflon film by laser ablation using a carbon dioxide laser. Multiple lipid membranes could be formed across the micro structured 8 × 8 array ETFE partitions. Success rates for the establishment of cellulose-supported BLMs across the multiple aperture arrays were above 95%. However, the time course of the membrane thinning process was found to vary considerably between multiple aperture bilayer experiments. An airbrush partition pretreatment technique was developed to increase the reproducibility of the multiple lipid bilayers formation during the time course from the establishment of the lipid membranes to the formation of bilayers. The results showed that multiple lipid bilayers could be reproducible formed across the airbrush-pretreated 8 × 8 rectangular arrays. The ionophoric peptide valinomycin was incorporated into established membrane arrays, resulting in ionic currents that could be effectively blocked by tetraethylammonium. This shows that functional bimolecular lipid membranes were established, and furthermore outlines that the established lipid membrane arrays could host functional membrane-spanning molecules.
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