Single Cell Isolation Via Microfluidic-Based Droplet Delivery

Commercial technologies such as fluorescent-activated cell sorting (FACS) for isolating and delivering single cell samples are often inappropriate for smaller research facilities due to their size and cost.  Researcher labs conducting experiments on single cells require an inexpensive and accessible method for creating samples.   We demonstrate the practicability of using a gravity-driven microfluidic device under a bench-top optical trap to generate droplets containing isolated single cells for delivery onto a 96 well plate.  The microfluidic device dispenses droplets of constant volume at fixed intervals.  We establish functionality by delivering droplets containing glutaraldehyde fixed red blood cells (RBCs) onto glass slides.   A fluorescent confocal microscope capable of imaging the glutaraldehyde-induced fluorescence of the RBCs is employed to count the number of cells contained per droplet.   We attained 64% fidelity of single-cell samples, indicating that delivery from our device is not stochastic. Biomedical researchers often expect droplet delivery methods to be compatible with a 96 well plate standard; to this end we achieved automated delivery into each well of a 96 well plate using a Teensy 3.0 encoded motorized stage.  Additionally, we implemented a side and forward scatter detection scheme around the optical trap to show the feasibility of fully automating the cell isolation and droplet delivery process.   The entire apparatus is compact and portable enough to fit within a sterile environment to prevent contamination of samples.  We propose that our setup is an inexpensive and compact alternative to flow cytometers that is accessible to most labs and can reliably sort single cells into droplets for sterile delivery.