Rapid Hemoglobin F and F-Cell Quantitation

The goal of this project is to develop two simple sensitive and accurate immunologic methodsto quantify hemoglobin F. One method will enumerate and identify the erythrocytes and reticulocytescontaining hemoglobin F and the other would quantify the percentage of hemoglobin F in bloodhemolysates. The development of these kits is facilitated in this laboratory by the availability ofmonoclonal antibodies against hemoglobin F. This obviously permits the development of better methodsthan the cytologic acid elution method of Kleihauer or the alkaline precipitation methods of Betke. Thus,using available antibody, the Investigator plans to develop a enzyme link immunoabsorbent assay (ELISA)to quantify the absolute level of hemoglobin F in red blood cell lysates and secondly to develop afluorochrome conjugated immunoassay to specifically enumerate the intact red blood cells which producehemoglobin F. The kits will be developed which could use either flow cytometry and/or fluorescencemicroscopy to enumerate hemoglobin F cells in the fluorochrome conjugated immunoassay. The lattermethodology would permit measurements of the relative proportion of reticulocytes containing Hb F ifdual staining was used or studies of the nature of the other Hb molecules which share the sameerythrocytes with Hb F and the relative amounts of Hb F within individual cells. A simple assay suchas those described would be highly useful in drug studies developed to increase hemoglobin F synthesisto ameliorate the severity of sickle cell disease.Likewise, it may be useful in studies which were desiredto determine the number of fetal nucleated red cells that entered the maternal circulation.