A cross-sectional study of 269 medical first responders (MFRs) in the fifth public health region of Thailand was conducted by voluntary participation between February and August 2011 to assess personal health risk histories, occupational risk contacts, and preventive practices towards hepatitis B virus (HBV) infection. Data from all voluntary MFRs were collected using a structured questionnaire consisting of socio-demographic characteristics, personal health risk histories, occupational risk contacts, and preventive practices towards HBV infection. Results revealed that 83.6% of studied MFRs were male, 88.8% finished only secondary level or lower, and 29.7% did not pass the training before working. Approximately 62.1% used to tattooing and/or piercing a hole in the ears, nose and other parts of the body, 7.8% had a history of injecting drug use, and 28.6% had a history of extramarital sex relations without using condom. For occupational risk contacts, 65.4% had a history of blood and/or secretion contact, and 42.4% had a history of needle stick and/or sharp injury during working. Their preventive practice mean score towards HBV infection showed rather low level with the mean score of 4.19±2.11 from a total score of 7. Therefore, the MFRs should be provided with training in universal precaution practices and life-skill education in order to reduce the risk of HBV infection during working.