Analytical Platform for multiscale biosensing on 2D pores networks – PLANARPORE

Our health depends in many cases of the course of our immune response. It organizes the body's defenses against infections, is involved in the mechanisms of inflammatory and autoimmune diseases, the elimination of cancer cells, transplant rejection ... The precise definition of the functions of immune cells, particularly of their secretions, is of interest for both better diagnosis and adjustment of medicines within immunotherapies. Among these cells, lymphocytes play a key role, particularly through the production of cytokines (destruction of infected cells, antibody production ...). These monodisperse and non-adherent cells are besides suitable for handling in a microfluidic system. Biosensors can enable their study at the scale of the individual cell but multi-scale analytical systems for the study, within the same device, of biological objects of different sizes such as one cell (about 10 microns) and its secreted proteins (nanometric) are still research concerns. The challenge to make these analytic microdevices efficient is in the development of their architecture which must include multistage detection for cells and secretions. For twenty years, the use of functionalized pores has emerged as a method for biosensing reaching higher sensitivities than conventional methods. It is therefore suitable for the serial detection of very small amounts of samples. In this project, we propose to develop multiscale biosensors based on a network of pores i) of different sizes (10-100 microns and 10-200 nm) and ii) functionalized with recognition biomolecules for the specific capture of lymphocytes and the detection of their secretions. We have developed in 2009 a novel technique called CLEF (ContactLess ElectroFunctionalization) which allows fast, one-step and very localized grafting of biomolecules on the walls of a perforated pore (micro and nanopores) in a semiconductor material coated with a dielectric layer. However, these "through" pores, because of their 3D configuration, i) have a great complexity of manufacture and implementation, ii) do not allow further analyses using pores positioned in series for cascade detections, iii) are not compatible with standard optical detection techniques, iv) do not allow parallelization of the analysis and a multiplexing of the measurement. To overcome these limitations, we propose to use a two-dimensional pore technology based on the restriction of a microfluidic channel, named 2D pore, and to demonstrate the feasibility of CLEF for their localized functionalization. 2D pores will be designed using lithographic techniques on a microfluidic chip and CLEF will be implemented using microelectrodes placed in the fluidic channels close to the pores. The design of multiscale biosensors will then be envisioned with different sized pores successions for the combined detection of lymphocytes and their secretions. The microelectrodes located in the fluidic channels will allow the electrical detection of secretions of lymphocytes in nanopores. Biochemical interactions in each pore can also be detected by fluorescence microscopy due to the easy optical observation permitted by the glass covering of the chip. The 2D format of this multi-scale sensor allows to consider parallelized analysis to simultaneously process a large number of samples. This generic biosensor for dynamic multi-scale detection can adapt to any type of cell / secretions and bacteria / toxins biological model, and thus cover a wide range of healthcare applications.