Screening for hemoglobins S and C in newborn and adult blood with a monoclonal antibody in an ELISA procedure

Summary To facilitate the screening of blood for the presence of hemoglobins S or C, we devised an enzymelinked immunoassay (ELISA). The ELISA procedure incorporated a murine monoclonal antibody (mAb), ^S-1, which recognized both Hb variants but did not react with Hb A, Hb A₂ or Hb F. Hemoglobins in cord or adult hemolysates were coated on the surface of wells of polystyrene microtiter plates and treated with ^S-1 mAb, followed by goat anti-mouse IgG conjugated with horseradish peroxidase. After addition of tetramethylbenzidine substrate solution, a deep blue color developed, signifying the presence of Hb S or Hb C. The ^S-1 mAb ascites fluid could detect purified Hb S and Hb C when diluted to over 1/512,000 and cord blood hemolysates containing Hb S or Hb C when diluted to 1/128,000. Although maximal reactivity was achieved using undiluted hemolysates, the ELISA system could easily detect Hb S and Hb C in cord blood hemolysates when diluted 10^–4. The sensitivity of the ELISA was 1%, which exceeds the lowest quantities of these variants normally found in cord blood. In addition, we found that the ELISA procedure was suitable for detecting Hb S/Hb C in whole blood as well. The entire assay could be conducted on multiple samples in less than 1 h, thus providing a specific, sensitive, rapid and simple screening technique for Hb S and Hb C in cord or adult blood.Abbreviations ELISA enzyme linked immunoassay - mAb monoclonal antibody - Hb hemoglobin(s) - TBS tris-buffered saline - PBS phosphate-buffered saline - BSA bovine serum albumin