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α0-Thalassemia occurs from a deletion of 2 linked α-globin genes and interaction of these defective genes leads to hemoglobin (Hb) Bart's hydrops fetalis, the most severe and lethal thalassemia syndrome. Identification of α0-thalassemia carriers is thus essential for the prevention and control program. An immunochromatographic (IC) strip test was developed for rapid screening of α0-thalassemia by testing for Hb Bart's in the blood samples using a specific monoclonal antibody against Hb Bart's. To evaluate its sensitivity and specificity, the IC strip test was assessed in a cohort with various thalassemia genotypes from 4 different laboratories in Thailand and Australia. The result showed 97% sensitivity in α-thalassemia carriers with 2 α-globin genes deletion and Hb H disease. This is, in particular, the useful rapid screening test for regions where β-thalassemia and homozygous Hb E are also common. Similar hematologic and Hb data make it impossible to address the concomitant inheritance of α0-thalassemia in these samples without polymerase chain reaction (PCR)-based techniques, leading to misdiagnosis of the risk of having Hb Bart's hydrops fetalis. However, α-globin genotyping should be carried out in samples with positive IC strip as positive reactivity was also observed in homozygous α+-thalassemia carriers who have 2transα-globin gene deletions. These results indicate that in combination with red blood cell indices, the IC strip test could rule out mass populations for further α0-thalassemia detection by PCR-based analysis. The Alpha Thal IC strip also has the potential to replace testing for Hb H inclusion bodies, as it appears to be more sensitive, specific, and less labor intensive.