Background: Healthcare professional, is an individual who provides preventive, curative, promotional or rehabilitative health care services to people, families or communities. Exposure to blood borne pathogens (HIV, hepatitis B, hepatitis C) poses a serious risk to them. Education is a major component of prevention strategies to reduce the risk of occupational transmission of blood-borne pathogens to health care personnel. Web based learning is increasingly being adopted in health professionals' education. Therefore, this study aimed at assesses the effect of Web-based Course on Knowledge and Attitude of Health care Professionals in relation to Blood borne Pathogens. It has been carried out using a quasi-experimental research design. The subjects were constituted of 62 health-care professionals including (doctor, nurses, dentist, and lab technicians who have bachelor degree and higher. In order to collect the necessary information for this study; online self- administered questionnaires was used. There was a statistical significant difference in mean scores of knowledge, and attitude of health-care professionals pre and post web-based blood borne course (P<0.05). Conclusion: it is concluded that, web based leaning can effectively improve healthcare professionals' knowledge, and attitude toward blood-borne pathogens.