Evaluation of red cell membrane cytoskeletal disorders using a flow cytometric method in south of Iran Print

The diagnosis of hereditary red blood cell (RBC) membrane disorders, in particular hereditary spherocytosis (HS), and SAO is based on clinical history and RBC morphology and other conventional tests such as osmotic fragility. However, there are some milder cases of these disorders which are difficult to be diagnosed. The application of eosin-5'-maleimide (EMA) was evaluated for screening of RBC membrane defect along with some other anemia. We used EMA dye which binds mostly to band 3 protein and in lesser extent to some other membrane proteins for screening of some membrane defects such as HS. Fresh RBCs from hematologically normal controls and also patients with HS, SAO, hereditary elliptocytosis, Hereditary spherocytosis with pincered cells, severe iron deficiency, thalassemia minor, and autoimmune hemolytic anemia were stained with EMA dye, and analyzed for mean fluorescent intensity(MFI) using a flow cytometer. Results and RBCs from patients with HS and iron deficiency showed a significant reduction in MFI compared to those from normal controls (ρ<0.0001 and ρ<0.001 respectively), while macrocytic RBCs showed a significant increase in MFI (ρ<0.01). A significant correlation was shown between MCV and MFI, which the only exceptions were HS and thalassemia minor. Our results showed that the flow cytometric method could be a reliable screening and confirming diagnostic method with high sensitivity and specificity (95% and 93%, respectively) than conventional routine tests for HS patients prior further specific membrane protein molecular tests.