Alginate scaffold with a three-dimensional (3D) porous structure can provide sufficient space for the cell to adhere, and has a good biocompatibility and mechanical strength. In this work, low-intensity pulsed ultrasound (LIPUS) is used to enhance the porosity of alginate scaffold based on acoustic cavitation. In the experiment, the alginate-calcium-based 3D scaffold culture system is fabricated with an optimum CaCl2/sodium alginate ratio of 3:5. The mechanical properties of alginate scaffold are measured and scanning electron microscopy is used to analyze the porosity of the scaffold. In addition, the microscopy observation of green fluorescent protein expression and the CCk-8 assessment are adapted to analyze the cell proliferation effect. Experimental results show that with LIPUS treatment under appropriate driving parameters (acoustic pressure 0.055 MPa and treatment time 20 min), the porosity of the 3D scaffold can be significantly improved, which would benefit the cell growth in the scaffold.