Nanomanufacturing technology plays a crucial role in advancing sophisticated biomedical devices, biochips, tissue engineering, and advanced biomedical materials. Two-photon polymerization (TPP) offers nanoscale fabrication precision, eliminates the need for masks, and allows the creation of arbitrary three-dimensional structures, providing technical advantages that are unparalleled by traditional methods. The application of TPP technology in the biomedical field presents new challenges related to materials and systems. Although there has been significant discussion regarding biomaterials, comparatively little attention has been given to the limitations of manufacturing systems for biomedical functional devices. Currently, commercial TPP systems predominantly rely on point-by-point scanning for fabrication, which leads to low throughput. From a biomedical perspective, the goal is to achieve manufacturing precision at the single-cell level while scaling production throughput to the organ level. To expand the applications of TPP in biomedical engineering, advancements in both precision and throughput are critical. This review begins by introducing the fundamental principles of TPP. It summarizes recent advancements in the applications of TPP within the fields of tissue engineering, medical devices, and microfluidics. It then delves into the technological progress of TPP in recent years, focusing on aspects such as system design, manufacturing processes, and fabrication principles. The review highlights advancements in areas like the kinetics of light-matter interactions and the development of cutting-edge techniques such as spatiotemporal focusing. Finally, it discusses the future development directions of TPP technology in the biomedical field.