Real-time imaging of non-adherent cells maintained in contact with adherent cell layers

Abstract

Abstract Cell-cell interactions are crucial to the survival and function of cells. Imaging these interactions in real-time is a challenging task with non-adherent cells due to their mobility in suspension, not to mention their communication with other heterotypic cells. Here we present a novel method to trap non-adherent cells in precise locations with a layer of adherent cells in order to study and image their interactions while in contact over time. This platform uses a well plate format to integrate 8 individual microfluidic units. Each unit has a 3 mm × 3 mm × 40 μm (L × W × H) chamber for cell culture, and an array of one hundred 72 × 72 × 15 μm (L × W × H) trap areas within each chamber to immobilize non-adherent cells underneath. The chamber in each unit can be addressed with programmable and on-demand perturbation of up to 2 different reagents to perform live cell imaging assays without disrupting the cell cultures. We have successfully cultured MCF-7 cells using this design, with Jurkat cells subsequently loaded and trapped above the layer of MCF-7. Upon reagent introduction, time-lapse images confirmed that the Jurkat cells remained in place. In conclusion, the microfluidic design allows investigation of cell-cell interactions in a wide range of applications requiring the confinement of non-adherent cells in contact with a different adherent cell layer. The on-demand perturbation feature further increases the capability to control the microenvironment exposed to the cell culture.