Characterization of Switching Time and Cell Stress in a Gravity-Driven Microfluidic Cell Sorter Based on Hydrodynamic Switching

In this work we describe the control and characterization of the switching time and hydrodynamic stress in a microfluidic cell sorter. The device was designed to sort small (<1000) populations of live cells in buffer solution labeled with standard bio-markers such as live dyes or green fluorescent protein (GFP). Sorting occurs through a hydrodynamic switching technique where high-speed solenoid valves control a sheath flow used to steer sorted cells away from the unsorted bulk population. The device is constructed from a reusable hard plastic polymethylmethacrylate (PMMA) chip machined with 127μm × 50μm microchannels and sealed with adhesive tape. Open reservoirs in the chip facilitate pipette access, standard microscope visualization, and a simple disassembly and cleaning procedure. The sorting frequency of this type of device is typically limited by the hydrodynamic switching time. Here, we present a theoretical and numerical analysis of the device switching time. These results show that the sorter switching time t is practically limited by the velocity of the flow and the characteristic length between inlet and outlet channels. We validate this theoretical result with experimental data obtained from flow visualizations, along with experiments conducted to evaluate the repeatability of the hydrodynamic switching scheme and the survival rate of sorted fibroblast cells Manually operated, the sorting frequencies were approximately 10 cells per minute, with switching time constants of approximately 130ms. Current throughput is limited by this switching time to approximately 450 cells per minute. Automation can increase the velocity and reduce the spacing between cells, thereby increasing throughput by at least an order of magnitude. The cell sorter was then tested by manually sorting 100 beads in 7 minutes, and 30 cells in less than 3 minutes, and was successfully used in the framework of a study on the bystander effect occurring during cell irradiation. Experiments with Trypan Blue dye verified that cell viability was maintained during the sorting process.