Hepatitis B exposure was assessed in 311 health workers in Uganda, a highly endemic country. Health workers were selected by random sampling from a categorized list of health workers at district level, proportionate to the population of each district. Whereas 60.1% of health workers have evidence of hepatitis B infection, with 8.7% being chronic carriers and one (0.3%) acutely infected, 36.3% are still susceptible and could benefit from vaccination. Only 5.1% reported having had at least one dose of hepatitis B vaccine and 3.5% were apparently immune through vaccination. Needle stick injuries reported by 77% of health workers were the most common mode of exposure to blood and body fluids. Trends suggested duration of service as a predictor while age and history of blood transfusion remained significant independent risk factors for hepatitis B infection. 98% of health workers are willing to be vaccinated. These results confirm the need for protection and vaccination of health workers in Uganda against hepatitis B.