The rapid development of medical care innovations and the use of newer technologies have resulted in a significant rise in the quantity of waste produced per patient within health care facilities. Biomedical waste in particular has emerged as a pressing concern due to its inherent propensity to pose health hazards and cause environmental harm. The World Health Organization has identified a lack of knowledge and training as primary factors contributing to the failure of Biomedical waste management (BMWM). Thus, our study sought to examine the relationship between BMWM level of knowledge and prior training among health care professionals (HCPs) in 4 different regions in Saudi Arabia. Our study used a cross-sectional design to investigate the BMWM level of knowledge and prior training among HCPs in 4 different regions in Saudi Arabia. Multiple linear and logistic regression was used to assess if Biomedical waste management (MBWM) knowledge and prior tanning are significantly associated with the demographics and professional characteristics of HCPs. 501 HCPs participated in the study. More than half were male, the majority fell within the age range of 24 to 36 years and held a bachelor's degree. The level of BMWM knowledge among HCPs was moderate (M=31.14, SD=4.89) and positively associated with HCPs' prior training in BMWM, workplace environment, and overall experience in the field. Only 26% of the HCPs had received prior BMWM training, and it varied considerably by sex, geographic location, and professional experience. Our study demonstrates a moderate knowledge and lack of training in BMWM among HCPs in Saudi Arabia. Thus, we recommend a national strategy for incorporating infection prevention and control methods associated with BMWM into all health care programs. Furthermore, it is imperative for health officials to enact prompt and efficient surveillance measures, as well as conduct frequent training sessions for HCPs and support personnel.