Abstract Perovskite solar cells (PSCs) show great potential for next-generation photovoltaics, due to their excellent optical and electrical properties. However, defects existing inside the perovskite film impair both the performance and stability of the device. Uncoordinated Pb2+, uncoordinated I−, and metallic Pb (Pb0) are the main defects occur during perovskite film preparation and device operation, due to the volatilization of organic cationic components. Passivating these defects is a desirable task, because they are non-radiative recombination centers that cause open-circuit voltage (VOC) loss and degradation of the perovskite layer. Herein, the multifunctional bioactive compound dopamine (DA) is introduced for the first time to control the perovskite film formation and passivate the uncoordinated Pb2+ defects via Lewis acid-base interactions. The Pb0 and I− defects are effectively suppressed by the DA treatment. At the same time, the DA treatment results in a stronger crystal orientation along the (110) plane and upshifts the valence band of perovskite closer to the highest occupied molecular orbital (HOMO) of the hole transport layer (2,2′,7,7′-tetrakis(N,N′-di-pmethoxyphenylamine)-9,9′-spirobifluorene, spiro-OMeTAD), which is beneficial for charge separation and transport processes. Consequently, the stability of MAPbI3 (MA = CH3NH3) PSCs prepared with the DA additive (especially the thermal stability) is effectively improved due to the better crystallinity and lower number of defect trap states of the perovskite film. The optimized MAPbI3 PSCs maintain approximately 90% of their original power conversion efficiency (PCE) upon annealing at 85 °C for 120 h. The best performance triple-cation perovskite (Cs0.05(FA0.83MA0.17)0.95Pb(I0.83Br0.17)3) (FA = formamidinium) solar cell with ITO/SnO2/Cs0.05(FA0.83MA0.17)0.95Pb(I0.83Br0.17)3:DA/spiro-OMeTAD/MoO3/Ag (ITO = indium tin oxide) structure shows a PCE of 21.03% with negligible hysteresis, which is dramatically enhanced compared to that of the control device (18.31%). Therefore, this work presents a simple and effective way to improve the efficiency and stability of PSCs by DA treatment.