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The rapid developments in biofabrication, in particular 3D bioprinting, in the recent years have facilitated the need for novel biomaterials that aim to replicate the target tissue in great detail. The presence of endotoxins in these biomaterials is often an overlooked problem. In pre-clinical 3D in vitro models, endotoxins can have significant influence on cell behavior and credibility of the model. In this study we demonstrate the effects of high levels of endotoxins in commercially-available gelatin on the macrophage-cancer cell crosstalk in a 3D bioprinted co-culture model. First, it is demonstrated that, while presenting the same mechanical and structural stimuli, high levels of endotoxin can have significant influence on the metabolic activity of macrophages and cancer cells. Furthermore, this study shows that high endotoxin contamination causes a strong inflammatory reaction in macrophages and significantly inhibits the …