Introduction Healthcare professionals continue to be at risk of acquiring occupation-related hepatitis B virus infection because of noncompliance for the 3-dose primary series of hepatitis B vaccine recommended. The objective of the study was to determine the rate of and to identify the predictors of hepatitis B vaccination uptake in healthcare professionals in Ethiopia. Methods A multicenter cross-sectional survey was conducted between May 19 2018 and June 15 2018. A stratified with systematic random sampling technique was used to select 260 healthcare professionals. A structured questionnaire was used to collect all the necessary primary data from samples. This survey analyzed hepatitis B vaccination uptake as the binary outcome variable (“noncomplete” vs “complete”) with regard to the 30 potential predictor variables. Both descriptive and inferential statistical analysis techniques have been used to address the study objective. Results The rate of complete vs noncomplete hepatitis B vaccination uptake was 37.7% vs 62.3%. The significantly independent predictors of hepatitis B vaccination uptake were professional qualification ( P =0.004), professional experience ( P =0.013), household income ( P =0.006), workload status ( P =0.015), routine infant immunization program existence at the workplace ( P =0.014), perceived susceptibility to infection ( P =0.005), perceived safety of vaccine ( P =0.001), prior occupational exposure to blood ( P =0.006), training on universal precautions ( P =0.015), and colleagues’ suggestion ( P =0.002). Conclusion The rate of hepatitis B vaccination uptake found was currently low. The significantly independent predictors of hepatitis B vaccination uptake included perceived safety, colleagues’ suggestion, professional qualification, perceived susceptibility, household income, prior occupational exposure to blood, professional experience, existence of routine infant immunization program at the workplace facility, workload status, and universal precautions training.