Introduction: Organ donation rate in Malaysia is amongst the lowest in the World. Healthcare professionals (HCPs) working in critical care areas play an important role in the deceased organ donation (DOD) process. This study seeks to identify the demographics of HCPs working in the critical care areas and their knowledge and attitudes toward the DOD process. Method: A cross-sectional survey on the demographics, knowledge and attitudes of the doctors and nurses working in critical care areas was undertaken by the random sampling method, using a validated, structured questionnaire. HCP’s knowledge and attitudes towards brain death (BD), DOD, organ transplantation (OT), and possession of organ donor card were compared against their demographics. Results: Four hundred and twelve (72.9%) out of the total 565 HCPs in critical care areas responded of whom 163 (39.6%) were doctors and 249 (60.4%) were nurses. After adjusting for other factors, department of work and profession were highly correlated with the overall knowledge score (p<0.001 and p=0.003 respectively) and knowledge about BD (p<0.001 and p=0.013 respectively). HCPs from the neurosurgical intensive care unit (p<0.001) and doctors (p<0.001) had higher mean knowledge scores compared to their counterparts. Profession was most significantly correlated with having a positive attitude towards BD (p<0.001) and OT (p<0.001). Conclusion: Department, profession and ethnicity were the demographic characteristics that correlated with knowledge and attitudes of HCPs on organ donation. Efforts to improve DOD rates in Malaysia should include targeted interventions to address the knowledge and attitudes of HCPs working in critical care areas.