Combination of dielectrophoretic and hydrodynamic traps for the controlled contact between cells or beads and application to adhesion assays

Multicellular organisms require very well organized and finely balanced cell-cell communication, adhesion and coordination to ensure the organisms homeostasis. These functions rely on specialized receptors placed at the cells membrane whose binding to their ligand triggers a defined response. Malfunction of the receptors to properly trigger the appropriate response leads to imbalances in the organism. Our understanding of these pathologies relies on the capacity to study such interactions. However, the tools currently capable of placing two micro-sized objects in contact for a controlled amount of time and probe their state of adhesion require skilled staff and are low throughput. Additionally, controlled contact between cells is of specific interest for adoptive cell therapies. Indeed, the process of selecting patient T-cells with high antitumor activity is currently a long and very expensive process, limiting the access to this therapy despite their proven efficacy. In this context, a tool able to reliably pair T-lymphocytes with tumor cells and rapidly assess the specificity of the interaction could facilitate access to this kind of therapy.This thesis tackles these challenges by developing a microfluidic tool combining two means for the controlled trapping of micro-sized objects to control their physical contact and probe their state of adhesion. Because the forces derive from a different phenomenon for each object, the effects are orthogonal and allow an independent manipulation of the two particles. The first part of the thesis describes a novel method for the hydrodynamic trapping of beads and cells relying on superimposed levels of microfluidic channels interconnected by microfluidic vias. This type of hydrodynamic traps enables the immobilization of objects without influencing neighboring objects. This method is a cornerstone of the chip as the controlled contact between two objects and the assessment of their state of adhesion can only be reliable when forces acting on them do not have intercrossed effects. The trapping mechanism of cells is first characterized in single traps to define the frame of operation for safe cell manipulation. The trapping of beads and cells and their arrangement in large arrays is then shown to demonstrate the capabilities of the method.Second, a design of coplanar electrodes creating a 3D dielectrophoretic force field directing to a single point against the flow is proposed. The DEP traps are used in combination with a deviation system to control the number of objects directed to the traps. The device is used to demonstrate single object trapping and the creation of aggregates composed of a controlled number of cells from two populations.Finally, the two means are combined to provide a device capable of performing in flow interaction with time of contact controlled down to the second resolution. The functionality of the tool is then validated by performing an adhesion frequency assay. The extracted binding kinetics parameters are compared to literature showing good agreement with previously reported values. The potential of such a tool in immunotherapy applications is demonstrated by pairing human T-cell clones with cancer cells. The lifetime of the pairs is measured and the effect of TCR-pMHC bonds on the latter is demonstrated.The developed device and method pave the way to faster devices for high-throughput T-cell screening and applications in adoptive cell therapies.