Removing IgG antibodies from intact red cells: Comparison of acid and EDTA, heat, and chloroquine elution methods

BACKGROUND: To accurately phenotype red cells from patients with a positive direct antiglobulin test (DAT), nonlytic elution procedures were assessed for their ability to dissociate IgG from antibody-coated red cells without altering red cell antigen expression. STUDY DESIGN AND METHODS: Antibodies coating red cells that were sensitized in vivo (warm-reactive autoantibodies: 8 patients) or in vitro (42 alloantibodies) were eluted by using glycine-HCl and EDTA (acid/EDTA), heat (56 degrees C, 10 min), or chloroquine method. RESULTS: Acid/EDTA elution gave the best results, reducing DAT positivity to microscopic levels or rendering the DAT negative in 48 of 50 instances, whereas 4 samples remained resistant to heat elution and 24 to chloroquine. Standard DAT agglutination scores demonstrated that both acid/EDTA and heat elution were superior to the chloroquine method (p<0.0001). With the gel low-ionic-strength saline indirect antiglobulin test, acid/EDTA was superior to heat (p<0.001). Overall, acid/EDTA elution dissociated more antibodies than heat (p<0.0001), especially for Kell system (K, k, Kp(a), Kp(b)) alloantibodies. Common red cell antigens, other than Kell system antigens, were unaffected by acid/EDTA elution. In contrast, the expression of most blood group antigens was diminished after heat elution. However, it was possible to type red cell antigens by using gel low-ionic-strength saline indirect antiglobulin tests or tube agglutination methods. CONCLUSION: Although heat elution may be used on a limited basis, the acid/EDTA method appears to be the procedure of choice for typing red cells coated with warm-reactive IgG alloantibodies or autoantibodies.