Background: β-thalassaemia (β-Thal) is an inherited chronic haemolytic anaemia resulting from absent or low level of synthesis of β-globin chains of haemoglobin A in erythropoietic cells. The complement system is an important part of innate immune response that may be implicated in red blood cell (RBC) lysis. Mammalian cells are provided with surface bound complement regulatory proteins (MCRPs) that regulate the activation of complement cascade, thus protecting them from uncontrolled complement-mediated lysis. Objective is to evaluate the role of complement regulatory proteins (CD55, CD59, and CD35) on red blood cells, and to explain the pathogenesis of anaemia in β-thalassemia major. Methods: This case-control study enrolled 74 β thalassemia major patients who were compared with 40 age and sex matched controls. We performed expression of CD55, CD59, and CD35 on RBCs using flow cytometry. Results: CD55 levels of β-thalassemia major patients (79.78% ± 18.54%) were significantly decreased compared to healthy controls (99.45% ± 0.59%) (P < 0.001). CD59 levels of β-thalassemia major patients (97.76% ± 1.72%) were significantly lower than in the controls (99.75% ± 0.36%) (P < 0.001), also CD35 levels were significantly lower in the β-thalassemia major patients (4.30% ± 4.66%) than in the control group (19.40% ± 10.90%) (P < 0.001). Conclusion: β-thalassemia major patients suffer from increased haemolysis and a consequent increase in their demand for blood transfusion. Complement-mediated haemolysis was shown in our study by decreased expression of CD55, CD59, and CD35 in β-thalassemia major patients. This allows complement deposition on RBCs and enhances or accelerates their lysis.