Decellularization of whole organs, such as the kidney hold great promise in addressing donor shortage for transplantation. However, successful implantation of engineered whole kidney constructs has been challenged by the inability to maintain endothelial cell coverage of the vasculature matrix, resulting in excessive blood clots, loss of vascular patency, and cell death within the construct. In this study, we describe an endothelial cell seeding approach that permits effective coating of the vascular matrix of the decellularized porcine kidney scaffold using a combination of static and ramping perfusion cell seeding. Furthermore, conjugation of CD31 antibodies to the vascular matrix improved endothelial cell retention on the vasculatures, which enhanced vascular patency of the implanted scaffold. These results demonstrate that our endothelial cell seeding method combined with antibody conjugation improves endothelial cell attachment and retention leading to vascular patency of tissue-engineered whole kidney in vivo. INNOVATIONA major challenge in the long-term success of bioengineered whole organs using decellularization approach is maintaining vascular patency following implantation. As such, uniform endothelial cell coverage of vascular walls of the matrices prevents thrombosis and permits blood fl ow into the scaff old. Unfortunately, none of current methods is able to address this critical challenge. To this end, this study presents with an eff ective method to achieve functional re-endothelialization of the vasculatures within the porcine kidney scaff old using a combination of static and perfusion seeding techniques. More importantly, we successfully conjugated endothelial cell specifi c antibodies to the vascular walls of decellularized whole kidney scaff olds that render vasculature surfaces more "sticky" to enhance endothelial cell attachment and prevent cell detachment under physiological fl ow conditions. We demonstrate the eff ectiveness of antibody conjugation mediated re-endothelialization in vitro by cell detachment testing and in vivo through implantation.

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