Growing 3D cultures presents several distinct challenges despite their ability to provide realistic models. One primary challenge is that new variables concerning how tissues function and how pathological processes disturb cellular functions need to be carefully addressed during the process.

Additionally, 3D models are significantly more expensive and time-consuming than 2D cultures. This is largely because there are very few options for automation and limited reproducible applications available. Consequently, these cultures require careful planning and expert handling to manage their complexity, unlike the more straightforward 2D monolayers.