In recent years, Raman spectroscopy is being used more frequently in biomedical analyses, especially for in-situ nondestructive detection of target samples or tissues. Raman probe, as the essential detection component in Raman spectroscopy, is also being developed for better diversity and functionality. Since Raman signal is extremely weak and noise interference is common, the new design and production of Raman probe are also extremely critical. This review provides an in-depth introduction to the existing general design and construction of Raman probe for biomedical applications, including the selection of fiber and filter/membranes, the design of probe tip, and the optimization of probe backend. Subsequently, the expansion and application of the Raman probe technology in the biomedical field are reviewed. The probes based on a combination of Raman and other spectra or imaging modes bring new vitality and prospects for broad applications in the biomedical and clinical detection of target samples or tissues.