AbstractIn vitro three-dimensional (3D) cell models have been accepted to better recapitulate aspects of in vivo organ environment than 2D cell culture. Currently, the production of these complex in vitro 3D cell models with multiple cell types and microenvironments remains challenging and prone to human error. Here we report a versatile bioink comprised of a 4-arm PEG based polymer with distal maleimide derivatives as the main ink component and a bis-thiol species as the activator that

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... ks the polymer to form the hydrogel in less than a second. The rapid gelation makes the polymer system compatible with 3D bioprinting. The ink is combined with a drop-on-demand 3D bioprinting platform consisting of eight independently addressable nozzles and high-throughput printing logic for creating complex 3D cell culture models. The combination of multiple nozzles and fast printing logic enables the rapid preparation of many complex 3D structures comprising multiple hydrogel environments in the one structure in a standard 96-well plate format. The platform compatibility for biological applications was validated using pancreatic ductal adenocarcinoma cancer (PDAC) cells with their phenotypic responses controlled by tuning the hydrogel microenvironment.