Background: End-stage renal disease (ESRD) has emerged as a major public health problem around the world. Infection is potentially preventable; yet, it remains the second most common cause of death in hemodialysis patients and still remains a major problem among patients on hemodialysis (HD) especially in Egypt. In recent decades, several important advances have been made in the therapy of HD with guidelines to ensure the delivery of optimum care to HD patients. Aim: to evaluate the impact of infection control measures as specified in guidelines on infection rate at hemodialysis unit. Study Design: a quasi-experimental study design was utilized to meet the aim of the study. Setting: It was conducted in the hemodialysis unit in Minya University hospital. Subjects: The study group was a convenience sample which included all health team staff in the hemodialysis unit. Data collection tools: 1) Self-Administered Questionnaire, 2) Infection Control Associated Practices Observational Checklists, 3) Polices and Resources' Assessment Tool and 4) Infection Assessment Tool for Patients in hemodialysis unit. Results: The study results revealed an obvious improvement in knowledge and practice of the studied health team staff in hemodialysis unit with a statistically significant differences pre, post and follow up the guidelines application. Also, the results revealed a decrease in signs and symptoms of infection and decrease in infection rate post guidelines application. Conclusion: Application of infection control guidelines improves health team staff knowledge and practices with positive impact on the infection rate for patients and health team staff in hemodialysis units. Recommendation: Continuous education and training of both new and current health team staff in hemodialysis unit about infection prevention and control guidelines should be enhanced.