Surface-tension driven open microfluidic platform for hanging droplet culture

The hanging droplet technique for three-dimensional tissue culture has been used for decades in biology labs, with the core technology remaining relatively unchanged. Recently microscale approaches have expanded the capabilities of the hanging droplet method, making it more user-friendly. We present a spontaneously driven, open hanging droplet culture platform to address many limitations of current platforms. Our platform makes use of two interconnected hanging droplet wells, a larger well where cells are cultured and a smaller well for user interface via a pipette. The two-well system results in lower shear stress in the culture well during fluid exchange, enabling shear sensitive or non-adherent cells to be cultured in a droplet. The ability to perform fluid exchanges in-droplet enables long-term culture, treatment, and characterization without disruption of the culture. The open well format of the platform was utilized to perform time-dependent coculture, enabling culture configurations with bone tissue scaffolds and cells grown in suspension. The open nature of the system allowed the direct addition or removal of tissue over the course of an experiment, manipulations that would be impractical in other microfluidic or hanging droplet culture platforms.