Monoclonal antibody-based methods for quantitation of hemoglobins: application to evaluating patients with sickle cell anemia treated with hydroxyurea.

High-titer monoclonal antibodies (mAb) were raised against chromatographically purified human hemoglobin (Hb) species. These mAb were specific for either Hb A, Hb F, Hb S or Hb C. Based on these antibodies, which were directly conjugated with either fluorochromes or an enzyme (horseradish peroxidase), we developed immunoassays for determining the Hb profile in the peripheral blood; an enzyme-linked immunosorbent assay (ELISA) for determining the absolute and relative quantities of various Hb species and one-step immunolabeling for fluorescence microscopic and flow cytometric analyses of the distribution of RBC with respect to their Hb types. We utilized these methods for monitoring the Hb F level and the percentage of Hb F-containing cells in patients with sickle cell anemia undergoing treatment with hydroxyurea.