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Three-dimensional cell spheroid cultures have become increasingly popular as drug screening platforms.One of the common methods for spheroid forming is the "hanging drop" technique,which offer an accessible and reliable method for generating higher numbers of cell spheroids with a precise and homogeneous size control and suitable compatibility for high throughput assays [1].Here,we present a simple and low-cost approach to fabricate hanging-drop system for spheroid culture.The hanging-drop system is based on a patterned superhydrophobic(SH)surface which is monolithically fabricated using a facile laser micromachining technique on regular polydimethylsiloxane(PDMS)substrate.By utilizing the wettability contrast,this system fixes liquid in the wettable regions to arrange arrays of quasi-spherical cell culture droplets with the capability to build-up 3D spheroids in a high throughput manner.Compared to the conventional microplate-based hanging-drop systems [2],this hanging-drop system produced by a benchtop methodology is more accessible and flexible.Figure 1 schematically outlines the direct-write laser machining process used to fabricate superhydrophobic patterns on a PDMS substrate.Firstly,the PDMS prepolymer was cast-coated on a clean glass slide.Subsequently,a CO2 laser machine directly engraved on the uncured coating polymer with predesigned patterns and control parameters.Finally,a post-processing bake was conducted to fully crosslink the uncured PDMS pre-polymer.After laser machining,the PDMS regions irradiated with the laser formed high-density nanofibrous structures(Figure 1b),exhibiting the superhydrophobicity,while the untreated regions remained wettable and transparent.Figure 2 shows examples of arrays of water microdroplets fixed on the patterned superhydrophobic surfaces.To demonstrate its ability to hanging drop culture,the developed platform was used as a support for arrays of quasi-spherical droplets of cell suspensions,for the generation of cell spheroids(Figure 3).We believe that such a platform may have the potential to be used as a new low cost toolbox for high-throughput drug screening and in cell or tissue engineering.