A chemiluminescent Langmuir–Blodgett membrane as the sensing layer for the reagentless monitoring of an immobilized enzyme activity

In the nanotechnology field, the concept of using biomolecules as an elementary structure to develop self-assembled entities has received considerable attention. Particularly, the ability of amphiphilic molecules like lipids to self-organize into bilayers can be exploited to provide biomimetic membrane models. Langmuir–Blodgett (LB) technology, based on the transfer of an interfacial film onto a solid support, offers the possibility to prepare lipid bilayers suitable for biomolecule immobilization and achievement of nanoscale-organized sensing layers, tailored for the design of miniaturized biosensors. With the aim of immobilizing enzymes in a defined orientation at the surface of LB bilayers, an original strategy has been previously developed in our group. This approach combines two techniques based on molecular self-assembly properties: liposome fusion at an air/buffer interface and Langmuir–Blodgett technology. It allows the functional insertion of a non-inhibitory antibody in lipid bilayers, further used to anchor a soluble enzyme at the surface of the lipid membrane. When associated with an electrochemiluminescent (ECL) sensor, this molecular assembly allows the design of a biomimetic sensor able to closely integrate the recognition and transduction events. However, sensorʹs performance not only depends on bioactive sensing layer properties, but also on the additional introduction of luminol in the reaction medium which delays ECL reaction. This work explores the potentiality of two neosynthesized amphiphilic luminol derivatives to form a lipid bilayer serving as a matrix used for both antibody insertion and ECL detection in order to develop a new sensing layer allowing a reagentless detection. As a model, choline oxidase activity has been detected. After enzyme immobilization at the surface of the luminol derivative LB bilayer by the way of specific recognition of a non-inhibitory antibody, in situ catalytic generation of hydrogen peroxide is able to trigger ECL reaction in the sensing layer interfaced with an optoelectronic device leading to a reagentless detection of choline oxidase activity.