Cellular spheroids, closely resembling native tissue microenvironments, have emerged as pivotal constructs in biomedicine as they can facilitate complex cell-cell and cell-matrix interactions. However, current methods for constructing spheroid assembloids with spatial arrangement or heterogeneous structures are limited, which has become a barrier for studying tissue engineering and in vitro disease modeling. Here, we demonstrate an acoustofluidic pick-and-place operation system capable of spatially assembling of spheroids into desired patterns in both two dimensional (2D) and three dimensional (3D) spaces. The underlying physical mechanism of the device is systematically studied to explain the interrelationship between trapping cell spheroids, acoustic streaming, and the acoustic radiation force (ARF) induced by the acoustically activated microneedle. We exploit these mechanisms to successfully transfer cellular spheroids into hydrogel solutions, enabling them to be precisely patterned and fused into assembloids of predefined shapes. Besides, we demonstrate arranging MC3T3-E1 cellular spheroids into a ring shape to fabricate the osteogenic tissues. Besides, a co-culture model involving tumor cells (MCF-7) and normal human dermal fibroblasts (NHDFs) is constructed to validate our method’s ability to reconstruct heterogeneous tumor model, revealing that the fibroblast spheroids promote tumor spheroid invasion. Our method holds significant potential prospects in regenerative medicine, disease model construction and drug screening.