Although it is quite challenging to image and analyze the spatial distribution of bioaerosols in a confined space, a three-dimensional (3D) modeling system based on the planar laser-induced fluorescence (PLIF) technique is proposed in this paper, which is designed to analyze the temporal and spatial variations of bioaerosol particles in a confined chamber. The system employs a continuous planar laser source to excite the fluoresce, and a scientific complementary metal oxide semiconductor (sCMOS) camera to capture images of 2048 × 2048 pixels at a frame rate of 12 Hz. While a sliding platform is moving back and forth on the track, a set of images are captured at different positions for 3D reconstruction. In this system, the 3D reconstruction is limited to a maximum measurement volume of about 50 cm × 29.7 cm × 42 cm, with a spatial resolution of about 0.58 mm × 0.82 mm × 8.33 mm, and a temporal resolution of 5 s. Experiments were carried out to detect the PLIF signals from fluorescein aerosols in the chamber, and then 3D reconstruction was used to visualize and analyze the diffusion of aerosol particles. The results prove that the system can be applied to clearly reconstruct the 3D distribution and record the diffusion process of aerosol particles in a confined space.