Background: Primary prevention of Monkeypox is achieved by public awareness campaigns that highlight the risk factors for the disease and the preventative steps that can be taken to lessen those risks, worry and stress are typical reactions to infectious illness epidemics such as Monkeypox. The public's uncertainty, concern, panic, and fear are understandable given that Monkeypox is a re-emerging disease that could cause severe consequences worldwide including increased concern and worry among mothers over their children's health and physical and mental well-being. Aim: To evaluate the effect of educational guidelines on mothers' knowledge, practice, and anxiety regarding Monkeypox. Design: To accomplish this study a quasi-experimental research design was used. Setting: This study was applied in the Pediatric Outpatient Clinic at Sohag University Hospital in Egypt. Sample: - A convenient sample of 200 mothers. Tools: Three tools were utilized: Tool I: Mothers' knowledge regarding Monkeypox, Tool II: Mothers' reported practice regarding Monkeypox, and Tool III: Zung's self-rating anxiety questionnaire. Results: The study findings revealed that mothers had a higher knowledge score and reported practice post-educational guidelines implementation than pre-educational guidelines implementation regarding Monkeypox. There was a decrease in the studied mothers' anxiety levels post-the educational guidelines implementation. There was a positive correlation between mothers' total knowledge, reported practice, and anxiety post-educational guidelines implementation.  Conclusion: According to the study's findings, mothers' knowledge, practice, and anxiety levels are all improved when educational guidelines about monkeypox are implemented. Recommendations: To assist and aid in psychological adjustment, it is crucial to offer mothers a well-thought-out health education program that will enhance their understanding and practice of monkeypox and reinforce potential interventions.