Background: Central line associated blood stream infections are serious but can often be prevented when evidence-based guidelines that are used by critical care nurse through utilized infection control measures during insertion and maintenance of central lines. Aim of the study was to evaluate the effect of educational guidelines on nurse's knowledge and practice regarding central line associated blood stream infection at intensive care unit. Subjects and Method: Research design: The present study utilized a quasi- experimental research design. Setting: The study was conducted at: Anesthesia Intensive Care Unit and Medical Intensive Care Unit of Emergency Hospital affiliated to Tanta Main University Hospital. Neurological Intensive Care Unit of Tanta Main University Hospital. The study subjects: All nurses (100) who are providing care for patients with central venous catheters and were having at least one year of experience. Tools: Two tools were used to collect the data; structured interviewing sheet and an observational checklist for nurses' practice. Results: The main results revealed that there was significant statistical improvement in the total knowledge and practice level among the studied nurses post nursing educational guidelines. Conclusion: the implementation of nursing educational guidelines has a positive impact not only in improving nurses' knowledge and practice regarding central line care but also, decrease rate of central line associated blood stream infection. Recommendation: it was recommended that nurses should attend in-service training programs for improving their knowledge and enhancing their practice