Sickle Hemoglobin: A Specific Radioimmunoassay

Abstract For the quantitation of hemoglobin S, a radioimmunoassay has been developed which is specific and highly sensitive. Hemoglobin S was purified by column chromatography and injected with complete Freund’s adjuvant into goats. Each goat serum was tested for reactivity against hemoglobins A and S by immunodiffusion and by quantitative precipitation. Hemoglobin A reactivity was removed by absorption with hemoglobin A. One serum so treated was specific for hemoglobin S; it reacted negligibly with hemoglobins A or F. Hemoglobin S was labeled with 125I by the chloramine T method. In the radioimmunoassay, complete precipitation of the antigen—antibody complex was insured by the addition of rabbit antigoat gamma globulin. This assay offers reliable and specific quantitation of as little as 1 ng of hemoglobin S. Assuming that 10% of the hemoglobin in fetal blood at 16 wk gestation is of adult type, this assay is capable of detecting the amount of hemoglobin S in 10-7 ml of homozygous hemoglobin S blood. The prenatal diagnosis of sickle cell anemia by radioimmunoassay will require, in addition, a method for demonstrating the absence of hemoglobin A and a safe method for obtaining fetal erythrocytes without significant contamination by maternal erythrocytes.