Abstract: Objective To understand the current status of construction and management of biosafety level-3 laboratories (BSLs-3) in China and provide evidence for public health policy-making. Methods The survey was conducted by using standard forms, the data collected were complied and analyzed. Results BSLs-3 were constructed or put into use in 40 institutions surveyed, The BSLs-3 which were put into formal use only accounted for 20%. The average construction fee of BSLs-3 was 6.464 million yuan RMB, and the average operation fee of the BSLs-3 was 384.75 thousand yuan RMB/year and the average maintenance fee was 225.7 thousand yuan RMB/year. The BSLs-3 were mainly used to detect 5 highly pathogenic microorganisms which frequently caused infectious diseases with high incidences in recent years. The main problems found in the construction and operation of BSLs-3 included the difficulty to pass the environment hazard evaluation and lack of operation fee. Conclusion The construction and management of BSLs-3 were standardized and legalized, but due to different construction condition and different understanding of related specifications, problems and difficulties still exist. It is suggested to strengthen the communication and cooperation among the related departments both at national level and local level to improve the construction and management of BSLs-3 in China.