Background: The aim of this study was to assess public and private medical diagnostic laboratories in Nigeria for the presence of biosafety equipment, devices, and measures. Methods: A total of 80 diagnostic laboratories in biosafety level 3 were assessed for the presence of biosafety equipment, devices, and compliance rate with biosafety practices. A detailed questionnaire and checklist was used to obtain the relevant information from enlisted laboratories. Results: The results showed the presence of an isolated unit for microbiological work, leak-proof working benches, self-closing doors, emergency exits, fire extinguisher(s), autoclaves, and hand washing sinks in 21.3%, 71.3%, 15.0%, 1.3%, 11.3%, 82.5%, and 67.5%, respectively, of all laboratories surveyed. It was observed that public diagnostic laboratories were significantly more likely to have an isolated unit for microbiological work (p = 0.001), hand washing sink (p = 0.003), and an autoclave ($p{\leq}0.001$) than private ones. Routine use of hand gloves, biosafety cabinet, and a first aid box was observed in 35.0%, 20.0%, and 2.5%, respectively, of all laboratories examined. Written standard operating procedures, biosafety manuals, and biohazard signs on door entrances were observed in 6.3%, 1.3%, and 3.8%, respectively, of all audited laboratories. No biosafety officer(s) or records of previous spills, or injuries and accidents, were observed in all diagnostic laboratories studied. Conclusion: In all laboratories (public and private) surveyed, marked deficiencies were observed in the area of administrative control responsible for implementing biosafety. Increased emphasis on provision of biosafety devices and compliance with standard codes of practices issued by relevant authorities is strongly advocated.