Pancreatic cancer, the fourth most common cause of cancer-related death, is a devastating disease with poor prognosis. Over the last four decades, no effective new treatments have been developed for this cancer. As a result, its prognosis has remained unchanged. Appropriate cancer models, representing all aspects of pancreatic cancer, will enhance our understanding of its biology. In this review we discuss the evolution and merit of organotypic culture models. These co-culture in vitro systems of cancer and stromal cells grown within, or on top of, reconstituted extracellular matrix gels model pancreatic cancer more realistically than 2D systems. Different methodologies are discussed which enable interrogation of various hypotheses examining the tumour-stroma cross-talk. Thus this validated organotypic culture model provides a system, which can be easily manipulated and used to test (novel) treatment options targeting the cancer, the stromal compartment or both, in a physiologically relevant environment. The big challenge for future research is to expand this model further so that its mimicry of the human tumour is more robust. This will increase our understanding of the biology of this aggressive tumour; ultimately resulting in improved therapies and, therefore, a better prognosis.