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Acoustofluidics, by combining acoustics and microfluidics, provides a unique means to manipulate cells and liquids for broad applications in biomedical sciences and translational medicine. However, it is challenging to standardize and maintain excellent performance of current acoustofluidic devices and systems due to a multiplicity of factors including device-to-device variation, manual operation, environmental factors, sample variability, etc. Herein, to address these challenges, we propose “intelligent acoustofluidics” – an automated system that involves acoustofluidic device design, sensor fusion, and intelligent controller integration. As a proof-of-concept, we developed intelligent acoustofluidics based mini-bioreactors for human brain organoid culture. Our mini-bioreactors consist of three components: (1) rotors for contact-free rotation via an acoustic spiral phase vortex approach, (2) a camera for real-time tracking of rotational actions, and (3) a reinforcement learning-based controller for closed-loop regulation of rotational manipulation. After training the reinforcement learning-based controller in simulation and experimental environments, our mini-bioreactors can achieve the automated rotation of rotors in well-plates. Importantly, our mini-bioreactors can enable excellent control over rotational mode, direction, and speed of rotors, regardless of fluctuations of rotor weight, liquid volume, and operating temperature. Moreover, we demonstrated our mini-bioreactors can stably maintain the rotational speed of organoids during long-term culture, and enhance neural differentiation and uniformity of organoids. Comparing with current acoustofluidics, our intelligent system has a superior performance in terms of automation, robustness, and accuracy, highlighting the potential of novel intelligent systems in microfluidic experimentation.