In vitro erythropoiesis of cord blood haematopoietic stem cells (HSCs) to produce fully enucleated red blood cells could provide an alternate resource for the erythrocyte. However, haematopoietic processes are highly complex and dynamic; defining process requirements to produce reproducible cells of high purity and yield is not an easy task. One major obstacle is the lack of knowledge in process characteristics. Design of experiments (DOE) is proposed as a tool to unveil process complexities that exist in HSC cultures. Characterisation and optimisation of in vitro erythropoiesis as a single-step culture is first performed via a simple DOE experimental strategy. The optimised DOE culture produced significantly better results (higher growth and faster maturation) than other single-step cultures. Subsequently, use of DOE to reveal in vitro process dynamics was attempted. This study was much more challenging and the repeatability of DOE process models was compromised in some cases. Process control of HSC culture bioprocesses is required for the delivery of reliable cell culture products suited for clinical applications. The availability of a convenient and economical online real-time process monitoring system can provide the means to translate stem cell culture bioprocesses from the bench-side into manufacturing production. The design and integration of such a system capable of simultaneous process monitoring of multiple analytes (ammonia, pH and oxygen) is presented. Operational and functional stability of this unique online real-time in situ monitoring platform was achieved. Stability of oxygen and ammonia sensors was achieved for up to three and six days respectively but biocompatibilities of both sensors require some improvements. Sensors of pH were biocompatible but their stability in cell culture is required.