<jats:p> <jats:bold> <jats:italic>Background—</jats:italic> </jats:bold> Tissue-engineered or decellularized heart valves have already been implanted in humans or are currently approaching the clinical setting. The aim of this study was to examine the migratory response of human monocytic cells toward decellularized porcine and human heart valves, a pivotal step in the early immunologic reaction. </jats:p> <jats:p> <jats:bold> <jats:italic>Methods and Results—</jats:italic> </jats:bold> Porcine and human pulmonary valve conduits were decellularized, and migration of U-937 monocytic cells toward extracted heart valve proteins was examined in a transmigration chamber in vitro. Homogenized tissue specimens were size fractionated by SDS-PAGE. The decellularization procedure effectively reduced the migration of human monocytes toward all heart valve tissue. However, only the antigen reduction of human pulmonary valves abolished the monocytic response (wall, 0.88±0.19% versus 30.20±3.93% migrated cells [mean±SEM]; cusps, 0.10±0.06% versus 10.24±1.83%) and was significantly lower ( <jats:italic>P</jats:italic> <0.05) than that of the decellularized porcine equivalent (wall, 5.03±0.14% versus 24.31±2.38%; cusps, 3.18±0.38% versus 10.24±1.83%). SDS-PAGE of the pulmonary heart valve tissue revealed that considerable amounts of proteins with different molecular weights that were not detected in the human equivalent remain in the decellularized porcine heart valve. </jats:p> <jats:p> <jats:bold> <jats:italic>Conclusions—</jats:italic> </jats:bold> We describe for the first time that the remaining potential of decellularized pulmonary heart valves to attract monocytic cells depends strongly on whether porcine or human scaffolds were used. These findings will have an important impact on further investigations in the field of heart valve tissue engineering. </jats:p>