Terrorist attacks are increasingly becoming more lethal and less discriminate. The threat of bioterrorism is increasing daily. The ease of production and the broad availability of biological agents and technical know-how have led to a further spread of biological weapons and an increased desire among nations as well as terrorists to have them. Health professionals in emergency departments are expected to play crucial roles in the management of victims of bioterrorism when bioterrorism occurs. This study explored the knowledge, attitudes, and preparedness of emergency department nurses and medical officers (MOs) toward potential bioterrorist attacks in Ghana. This qualitative study utilized focus group discussions and semistructured interviews to explore the knowledge, attitudes, and preparedness of emergency department nurses and MOs toward potential bioterrorist attacks in Ghana. Data were subjected to a qualitative content analysis in which three main thematic categories were developed. These thematic categories are as follows: (a) differences in bioterrorism knowledge between emergency department nurses and emergency department MOs, (b) unprepared emergency department nurses and MOs for care during bioterrorism attacks, and (c) positive attitudes of emergency department nurses and MOs toward bioterrorism preparedness. Although emergency MOs had better knowledge of bioterrorism than their nursing counterparts, both groups of health professionals were unprepared to respond to any form of bioterrorism. Both nurses and MOs indicated the need for staff education and infrastructure readiness to be able to respond effectively to a bioterrorist attack. A well-prepared emergency department and health professionals against bioterrorism could prevent high casualty rates in a bioterrorist attack and also serve a dual purpose of dealing with other natural disasters when they occur.