Natural biomolecules are often used to functionalize materials to achieve desired cell-material interactions. However, their applications can be limited owing to denaturation during the material functionalization process. Therefore, efforts have been made to develop synthetic ligands with polyvalence as alternatives to natural affinity biomolecules for the synthesis of functional materials and the control of cell-material interactions. This work was aimed at investigating the capability of a hydrogel functionalized with a novel polyvalent aptamer in inducing cell attachment in dynamic flow and releasing the attached cells in physiological conditions through a hybridization reaction. The results show that the polyvalent aptamer could induce cell attachment on the hydrogel in dynamic flow. Moreover, cell attachment on the hydrogel surface was significantly influenced by the value of shear stress. The cell density on the hydrogel was increased from 40 cells/mm(2) to nearly 700 cells/mm(2) when the shear stress was decreased from 0.05 to 0.005 Pa. After the attachment onto the hydrogel surface, approximately 95% of the cells could be triggered to detach within 20 min by using an oligonucleotide complementary sequence that displaced polyvalent aptamer strands from the hydrogel surface. While it was found that the cell activity was reduced, the live/dead staining results show that ≥98% of the detached cells were viable. Therefore, this work has suggested that the polyvalent aptamer is a promising synthetic ligand for the functionalization of materials for regulated cell attachment.