Universal precautions intend to prevent in the health care staff parenteral, mucus membrane, intact and nonintact skin exposure to pathogens by preventing their exposure to blood and body fluids from all patients as if they were all potentially infectious with blood-borne pathogens. The universal precautions practice applies the basic principle of infection control through hand washing, utilisation of appropriate protective barriers such as gloves, masks, gowns and eyewear to prevent contact with potentially infectious materials, and safe handling of sharps. Hands have been identified as the major source of germ transmission while providing health care. Every person involved in patient care, healthcare worker or otherwise, should be able to accurately perform hand hygiene and at the appropriate time. Keeping in view of this, the present study was conducted to assess the knowledge and practice of hand hygiene among healthcare workers This cross-sectional descriptive study was carried out on 50 healthcare workers in a healthcare setting. All participants had to complete a 25-item self-administered WHO hand hygiene questionnaire for health-care workers 31 for assessment of the knowledge and practice of hand hygiene. The level of hand hygiene knowledge was calculated by dividing the responses into three groups based on a score of more than 75% considered as good, 50-74% moderate, and less than 50% considered as low. Around 37% (n=18) of the respondents never received any formal training in hand hygiene. Only 14% (n=7) of the respondents had a good level of hand hygiene knowledge.