Heart valve disease (HVD) is a globally increasing problem and accounts for thousands of deaths yearly. Currently end-stage HVD can only be treated by total valve replacement, however with major drawbacks. To overcome the limitations of conventional substitutes, a new clinical approach based on cell colonization of artificially manufactured heart valves has been developed. Even though this attempt seems promising, a confluent and stable cell layer has not yet been achieved due to the high stresses present in this area of the human heart. This study describes a bioreactor with a new approach to cell conditioning of tissue engineered heart valves. The bioreactor provides a low pulsatile flow that grants the correct opening and closing of the valve without high shear stresses. The flow rate can be regulated allowing a steady and sensitive conditioning process. Furthermore, the correct functioning of the valve can be monitored by endoscope surveillance in real-time. The tubeless and modular design allows an accurate, simple and faultless assembly of the reactor in a laminar flow chamber. It can be concluded that the bioreactor provides a strong tool for dynamic pre-conditioning and monitoring of colonized heart valve prostheses physiologically exposed to shear stress.