Red blood cells (RBCs) transfusion is frequently used in the management of patients with chronic renal failure (CRF) undergoing hemodialysis and thalassemia magor. Consequently, they are subject to red cell alloimmunization. The purpose of this study was to detect the frequency of red blood cell alloantibodies and to determine types of these antibodies among multiple-transfused thalassemic and CRF patients. A total of 200 patients who received regular blood transfusion (150CRF & 50 thalassemia), from Beni Suef university hospital and general hospital in Beni Suef, were included in this study and control group of 50 individuals from Beni Suef university hospital out patient clinic were included in this study . Screening of antibodies was performed on patients' plasma samples and then antibodies were identified in patients' plasma that had positive antibody screening test using a panel of recognized blood group antigens. Ten patients (5%) demonstrated clinically significant alloantibodies (six (4%) CRF patients and four (8%) thalassemic patients). The most common alloantibodies involved were to Rh &Kell blood group systems. We concluded that Prophylactic transfusion of ABO, Rh and Kell phenotype-matched blood, for patients who are transfusion dependent (CRF & thalassemia) from the start of transfusion can be helpful in decreasing the rate of alloantibody synthesis.