Severe acute respiratory syndrome coronavirus 2 and other highly pathogenic pathogens need to be studied in the Biosafety Level 3 (BSL-3) laboratory, and it is difficult to obtain live viruses, which limits the development of scientific research and prevention. Pseudovirus is a recombinant virus with similar structure and biological characteristics to wild-type virus without the pathogenicity of live viruses. Pseudovirus can be studied in the BSL-2 laboratory, and it can be detected quantitatively and qualitatively by carried reporter gene. Pseudovirus has been widely used for detection of neutralizing antibody and evaluation of vaccines, receptor recognition, establishment of animal models, antiviral drug screening and quality control of nucleic acid detection etc. In order to better carry out detection of neutralizing antibody, evaluation of vaccine and drug screening etc. of SARS-CoV-2 and other new, emerging and severe infectious disease pathogens, this article reviews the research progress and application of pseudovirus technology.