Monoclonal Antibody to the y of Human Fetal Hemoglobin Used to Develop an Enzyme Immunoassay

A monoclonal antibody (mAb) that recognizes the y chain of human fetal hemoglobin(Hb F) has been produced by cell hybridization techniques. The mAb reacts with Hb F (a2y2), Hb Bail’s(y),and Hb Kenya (yhybrid), butdoes notcrossreactwith Hb A (a) or Hb A2 (a2&). We describe a direct enzyme-linked immunoassay (ELISA)for measurement of Hb F, in which hemoglobins from standards or from unknown hemolysatesare covalentlyboundto the wells of microtiter plates. The antigen is quantified by addition of the 7-specific mAb, followed by anti-mouse IgG conjugated with horseradish peroxidase, and incubationwith the substrate,tetramethylbenzldine. Absorbances at630 nm aredirectly proportional to the amount of Hb F present in the standards or samples. Results for Hb F in 53 hemolysates agreed well withvalues obtainedby “high-performance”liquidchromatography,RIA, alkali denaturation, and magnetic affinity immunoassay. This EUSA can detect a 0.5% proportion of Hb F in 1 h and offers distinct advantages over othertechniquescurrentlyin use.