Background: The waste produced in the course of health care activities carries a higher potential for infection and injury than any other of waste. Therefore it is essential to have safe and reliable method for its handling. Aims: Assess the knowledge regarding segregation of bio medical waste among class IV workers Methods and Materials: The present cross sectional descriptive study was conducted in General wards and ICU’s of Narayana Medical College hospital. 30 class IV workers were selected by non probability convenience sampling technique. Data was collected by using structured questionnaire. Analysis was performed by using descriptive and inferential statistics. Results: The results revealed that 50% of class IV workers are having Moderately Adequate knowledge followed by 43.43% are having inadequate knowledge and only 6.67% had adequate knowledge regarding segregation of bio medical waste in the hospital. Conclusion: The study findings concluded that majority of class IV workers had moderately adequate and inadequate knowledge regarding bio medical waste management. It indicates that there is a need to implement standard protocols and written policies regarding bio medical waste management and further need to disseminate the information to the health care team members through continuous education programs.