Adherence to biosafety and biosecurity standards and practices by those involved in the life sciences is essential to protecting the health of humans, animals, and the environment. Fostering a culture of responsibility is one of the underlying tenets of the Biological Weapons Convention, the Global Health Security Agenda, Joint External Evaluation of International Health Regulations, and the Global Partnership Against the Spread of Weapons and Materials of Mass Destruction. It also underpins national and international efforts to protect science from misuse and to protect workers and the public from laboratory incidents. Unlike the nuclear domain, there is no available guidance or unified perspective on the nature of this culture or how it may be assessed. The International Working Group on Strengthening the Culture of Biosafety, Biosecurity, and Responsible Conduct in the Life Sciences developed a framework to assess organizational culture in biological laboratories based on the model developed by the International Atomic Energy Agency for nuclear facilities. This framework encompasses 1) management systems; 2) behavior of leadership and personnel; 3) principles for guiding decisions and behaviors; and 4) beliefs, opinions and attitudes. Here we present an example of how the beliefs, opinions, and attitudes element was employed to assess the organizational culture of laboratorians from five regions in Mali. This is the first assessment using the framework and, as such, provides a model for others seeking to improve biosafety and biosecurity, and may ultimately lead to international harmonization of this concept and provide a pathway to strengthen Mali’s laboratory system.